Methods of Making and Using Therapeutically Active Thiophenepyrimidinone Compounds

ABSTRACT

Thiopheneprymidinone compounds useful in therapy, especially for use in the treatment and/or prevention of a steroid hormone dependent disorder, preferably a steroid hormone dependent disease or disorder requiring the inhibition of a 17β-hydroxysteroid dehydrogenase (17β-HSD) such as 17β-HSD type 1, type 2 or type 3 enzyme.

CROSS REFERENCE TO RELATED APPLICATION

This application is a division of co-pending application Ser. No.10/861,922, filed Jun. 7, 2004, now U.S. Pat. No. ______. Priority isclaimed from U.S. provisional patent application No. 60/477,017, filedJun. 10, 2003.

FIELD OF THE INVENTION

The present invention relates to novel thiophenepyrimidinone derivativeswhich represent inhibitory compounds of the 17β-hydroxysteroiddehydrogenase enzymes, preferably of the 17β-hydroxysteroiddehydrogenase type 1 (17β-HSD1), type 2 (17β-HSD2) or type 3 (17β-HSD3)enzyme, to their salts, to pharmaceutical preparations containing thesecompounds and to processes for the preparation of these compounds.Furthermore, the invention concerns the therapeutic use of saidthiophenepyrimidinone derivatives, particularly their use in thetreatment or prevention of steroid hormone dependent diseases ordisorders, such as steroid hormone dependent diseases or disordersrequiring the inhibition of 17β-hydroxysteroid dehydrogenase enzymes, inparticular 17β-HSD type I enzymes, and/or requiring the modulation ofthe endogenous 17β-estradiol and/or testosterone concentration.

BACKGROUND OF THE INVENTION

The publications and other materials used herein to illuminate thebackground of the invention, and in particular, cases to provideadditional details respecting the practice, are incorporated byreference.

Mammalian 17β-hydroxysteroid dehydrogenases (17β-HSDs) are NAD(H) orNADP(H) dependent enzymes which catalyze—besides other reactions—thefinal steps in male and female sex hormone biosynthesis. These enzymesconvert inactive 17-keto-steroids into their active 17β-hydroxy-forms orcatalyze the oxidation of the 17β-hydroxy-forms into the17-keto-steroids. Because both estrogens and androgens have the highestaffinity for their receptors in the 17β-hydroxy form, 17β-HSD enzymesplay an essential role in the tissue-selective regulation of theactivity of sex steroid hormones.

At present, 10 human members of the 17β-HSD enzyme family have beendescribed (types 1-5, 7, 8, 10, 11 and 12). The human 17β-HSD familymembers share less than 30% similarity in their primary structure. The17β-HSDs are expressed in distinct, though in some cases, overlappingpatterns. The different types of 17β-HSDs also differ in their substrateand cofactor specificities. In intact cells in culture, the 17β-HSDscatalyze the reaction in a unidirectional way: types 1, 3, 5 and 7 useNADP(H) as a cofactor and catalyze the reductive reaction (activation),while types 2, 4, 8 and 10 catalyze the oxidative reaction(inactivation) using NAD(H) as a cofactor. [see e.g. Labrie et al.(2000) Trends Endocrinol Metab., 11:421-7].

Due to their essential role in the tissue-selective regulation of theactivity of sex steroid hormones 17β-HSDs can be involved in theoccurrence and development of estrogen-sensitive pathologies (f. ex.breast, ovarian, uterine and endometrium cancers etc.) andandrogen-sensitive pathologies (f. ex. prostate cancer, benign prostatichyperplasia, acne, hirsutism, etc). Furthermore, many types of 17β-HSDhave been shown to be involved in the pathogenesis of particular humandisorders. For example, 17β-HSD3 is known to be involved in thedevelopment of pseudohermaphroditism, the 17β-HSD8 plays a role inpolycystic kidney disease and the 17β-HSD4 is related to the occurrenceof bifunctional enzyme deficiency. Therefore treatment of sexsteroid-sensitive diseases by administration of specific inhibitors ofthe 17β-HSDs enzymes have been suggested, optionally in combination withpotent and specific antiestrogens and antiandrogens [Labrie F et al.(1997) Steroids, 62:148-58].

Due to the fact that each type of 17β-HSD has a selective substrateaffinity, directional (reductive or oxidative) activity in intact cells,and a particular tissue distribution, the selectivity of drug actioncould be achieved by targeting a particular 17β-HSD isozyme. Byindividual modulation of the particular 17β-HSDs it is possible toinfluence or even control the local and paracrine concentration ofestrogens and androgens in different target tissues.

The best characterized member of the 17β-HSD family is the type 117β-HSD [EC 1.1.1.62]. This enzyme could be crystallized in differentstates of functionality (e.g. with and without ligand and/or co-factor).The 17β-HSD1 catalyzes in vitro the reduction as well as the oxidationbetween estrone (E1) and estradiol (E2). However, under physiological invivo conditions the enzyme only catalyzes the reductive reaction fromthe estrone (E1) to the estradiol (E2). The 17β-HSD1 was found to beexpressed in a variety of hormone-dependent tissues, e.g. placenta,mammary gland tissue or uterus and endometrium tissue, respectively.Estradiol itself is, especially in comparison to the significantly lessactive estrone, a very potent hormone, which regulates the expression ofa variety of genes by binding to the nuclear estrogen receptor and playsan essential role in the proliferation and differentiation of the targetcell. Physiological as well as pathological cell proliferations can beestradiol dependent. Especially many breast cancer cells are stimulatedby a locally raised estradiol concentration. Furthermore, the occurrenceor course of benign pathologies such as endometriosis, uterineleiomyomas (fibroids or myomas), adenomyosis, menorrhagia, metrorrhagiaand dysmenorrhea is dependent from the existence of significantly highestradiol levels.

Endometriosis is a well-known gynecological disorder that affects 10 to15% of women in the reproductive age. It is a benign disease defined asthe presence of viable endometrial gland and stroma cells outside theuterine cavity. It is most frequently found in the pelvic area. In womendeveloping endometriosis, the endometrial cells entering the peritonealcavity by retrograde menstruation (the most likely mechanism) have thecapacity to adhere to and invade the peritoneal lining, and are thenable to implant and grow. The implants respond to steroid hormones ofthe menstrual cycle in a similar way as the endometrium in the uterus.The infiltrating lesions and the blood from these lesions which areunable to leave the body cause inflammation of the surrounding tissue.The most common symptoms of endometriosis are dysmenorrhea, dyspareuniaand (chronic) abdominal pain. The occurrence of these symptoms is notrelated to the extent of the lesions. Some women with severeendometriosis are asymptomatic, while women with mild endometriosis mayhave severe pain. Endometriosis is found in up to 50% of the women withinfertility. However, currently no causal relation has been provenbetween mild endometriosis and infertility. Moderate to severeendometriosis can cause tubal damage and adhesions leading toinfertility. The aims of treatment of endometriosis are pain relief,resolution of the endometriotic tissue and restoration of fertility (ifdesired). The two common treatments are surgery or anti-inflammatoryand/or hormonal therapy or a combination thereof.

Uterine leiomyomas (fibroids or myomas), benign clonal tumours, arisefrom smooth muscle cells of the human uterus. They are clinicallyapparent in up to 25% of women and are the single, most commonindication for hysterectomy. They cause significant morbidity, includingprolonged and heavy menstrual bleeding, pelvic pressure and pain,urinary problems, and, in rare cases, reproductive dysfunction. Thepathophysiology of myomas is not well understood. Myomas are foundsubmucosally (beneath the endometrium), intramurally (within themyometrium) and subserosally (projecting out of the serosal compartmentof the uterus), but mostly are mixed forms of these 3 different types.The presence of estrogen receptors in leiomyoma cells has been studiedby Tamaya et al. [Tamaya et al. (1985) Acta Obstet Gynecol Scand.,64:307-9]. They have shown that the ratios of estrogen receptor comparedto progesterone and androgen receptor levels were higher in leiomyomasthan in the corresponding normal myometrium. Surgery has long been themain treatment for myomas. Furthermore, medical therapies that have beenproposed to treat myomas include administration of a variety of steroidssuch as the androgenic steroids danazol or gestrinone, GnRH agonists andprogestogens, whereby the administration is often associated a varietyof serious side-effects.

Everything that has been said above in relation to the treatment ofuterine leiomyomas and endometriosis equally applies to other benigngynecological disorders, notably adenomyosis, functional menorrhagia andmetrorrhagia. These benign gynecological disorders are all estrogensensitive and are treated in a comparable way as described herein beforein relation to uterine leiomyomas and endometriosis. The availablepharmaceutical treatments, however, suffer from the same majordrawbacks, i.e. they have to be discontinued once the side-effectsbecome more serious than the symptoms to be treated and symptomsreappear after discontinuation of the therapy.

Since the aforementioned malign and benign pathologies are all17β-estradiol dependent, a reduction of the endogenous 17β-estradiolconcentration in the respective tissue will result in an impaired orreduced proliferation of 17β-estradiol cells in said tissues. Therefore,it may be concluded that selective inhibitors of the 17β-HSD1 enzyme arewell suited for their use to impair endogenous productions of estrogens,in particular of 17β-estradiol, in myomas, endometriotic, adenomyoticand endometrial tissue. The application of a compound acting asselective inhibitor on the 17β-HSD1 which preferentially catalyzes thereductive reaction will result in a lowered intracellularestradiol-concentration since the reductive conversion of the estroneinto the active estradiol is reduced or suppressed. Therefore,reversible or even irreversible inhibitors of the 17β-HSD1 may play asignificant role in the prophylaxis and/or treatment of steroid-hormone,in particular 17β-estradiol, dependent disorders or diseases.Furthermore, the reversible or even irreversible inhibitors of the17β-HSD1 should have no or only pure antagonistic binding activities tothe estradiol receptor, in particular to the estrogen receptor αsubtype, since agonistic binding of the estrogen receptor would lead toactivation and therefore—by regulation of a variety of genes—to theproliferation and differentiation of the target cell. In contrast,antagonists of the estrogen receptor, so called anti-estrogens, bindcompetitively to the specific receptor protein thus preventing access ofendogenous estrogens to their specific binding site. At present it isdescribed in the literature that several malignant disease as breastcancer, prostate carcinoma, ovarian cancer, uterine cancer, endometrialcancer and endometrial hyperplasia may be treated by the administrationof a selective 17β-HSD1 inhibitor. Furthermore, a selective 17β-HSD1inhibitor may be useful for the prevention of the aforementionedhormone-dependent cancers, especially breast cancer.

Several reversible or irreversible inhibitors of the 17β-HSD1 enzyme ofsteroidal and even non-steroidal origin are already known from theliterature. The characteristics of these inhibitory molecules, whichmainly have a substrate or cofactor-like core structure, have beenreported in the literature [reviewed in: Poirier D. (2003) Curr Med.Chem. 10:453-77].

A further well characterized member of the 17β-HSD family is the 17β-HSDtype 3 enzyme (17β-HSD3). The 17β-HSD3 has a distinct feature comparedto other 17HSDs: it is found to be expressed almost exclusively thetestis, whereas the other isoenzymes are expressed more widely inseveral tissues. 17β-HSD3 has a crucial role in androgen biosynthesis.It converts 4-androstene-3,17-one (A) to testosterone (T). Thebiological significance of the 17β-HSD3 is of undeniable physiologicalimportance. Mutations in the gene for 17β-HSD3 have found to lead todecreased T formation in the fetal testis and consequently to a humanintersex disorder termed male pseudohermaphroditism [Geissler W M et al.(1994) Nat. Genet., 7:34-9].

With regard to the indication prostate cancer, the primary cancer cellsmostly retain their responsiveness to androgens in their regulation ofproliferation, differentiation, and programmed cell death for someperiod. At present, androgen deprivation is the only effective systemichormonal therapy available for prostate cancer. The development ofselective inhibitors against 17β-HSD3 is a new therapeutic approach forthe treatment of androgen dependent disease [Labrie et al. (2000) TrendsEndocrinol Metab., 11:421-7]. Furthermore, Oefelein et al. reported thatthe depot GnRH analogue fails, in nearly 20% of cases, to achievecastrate levels of T in men [Oefelein M G & Cornum R (2000) J Urol.;164:726-9]. In order to improve the response rate to endocrine therapyfor men with prostate cancer it may be important to selectively inhibittesticular 17β-HSD3 activity. Besides prostate cancer, many otherandrogen-sensitive diseases, i.e. diseases whose onset or progress isaided by androgenic activity, may be treated by selectively inhibiting17β-HSD3 activity. These diseases include but are not limited to benignprostatic hyperplasia, prostatitis, acne, seborrhea, hirsutism,androgenic alopecia, precocious puberty, adrenal hyperplasia, andpolycystic ovarian syndrome. Furthermore, considering the fact that17β-HSD3 is found mainly in the testis, the development of potentinhibitors could be of interest for blocking spermatogenesis and as ananti-fertility agent for males.

Several reversible or irreversible inhibitors of the 17β-HSD3 enzymes ofsteroidal and even non-steroidal origin are already known from theliterature. The characteristics of these inhibitory molecules have beenreported in the literature [reviewed in: Poirier D. (2003) Curr Med.Chem. 10:453-77]. For example, U.S. Pat. No. 6,541,463 disclosesandrosterone derived inhibitors for 17β-HSD3. These derivatives havebeen synthesized by parallel solid- and liquid-phase chemistry and someof these compounds showed 2 to 18-fold higher inhibition activity thanthat of the natural substrate of the enzyme, A-dione, used itself as ainhibitor. Furthermore, the international patent application WO 01/42181discloses benzyl-tetralins, the chemical structure of which is relatedto that of the phytoestrogen biochanin, as 17β-HSD3 inhibitors.Furthermore, international patent applications WO 98/32724, WO 98/30556and WO 99/12540 disclose tetralone, benzopyrane and benzofuranonederivatives, which have a 17β-HSD inhibitory activity, for the treatmentof hormone sensitive diseases.

Microsomal 17β-hydroxysteroid dehydrogenase of human endometrium andplacenta (designated 17β-HSD type 2 or 17β-HSD2) was cloned byexpression cloning, and found to be equally active using androgens andestrogens as substrates for oxidation [Andersson S. (1995) J. SteroidBiochem. Molec. Biol., 55:533-534]. The recombinant 17β-HSD2 convertsthe highly active 17β-hydroxysteroids such as estradiol (E2),testosterone (T), and dehydrotestosterone (DHT) to their inactive ketoforms. In addition, the 17β-HSD2 can, to a lesser extent, also convert20β-hydroxyprogesterone (20βP) to progesterone (P). The broad tissuedistribution together with the predominant oxidative activity of17β-HSD2 suggest that the enzyme may play an essential role in theinactivation of highly active 17β-hydroxysteroids, resulting indiminished sex hormone action in target tissues. Dong and colleaguesshowed significant 17β-HSD2 activity in cultured human osteoblasts andosteoblast-like osteosarcoma cells MG63 and TE85, but not in SaOS-2[Dong Y et al. (1998) J. Bone Min. Res., 13:1539-1546]. The potentialfor interconversion of E1 to E2, T to A, and DHT to A by bone cellscould therefore represent important mechanism for the local regulationof intracellular ligand supply for the estrogen and androgen receptorsin the osteoblasts and other steroid sensitive cells. This modulation ofsteroid levels may be employed for a wide variety of indications,including the following: for the prevention and treatment ofosteoporosis, for the treatment of ovarian cancer, for the treatment ofbreast cancer, for the treatment of endometrial cancer, for thetreatment of endometriosis, for the treatment of prostate cancer and/orfor the treatment of androgen-dependent hair-loss.

Several reversible or irreversible inhibitors of the 17β-HSD2 enzymes ofsteroidal and even non-steroidal origin are already known from theliterature. The characteristics of these inhibitory molecules have beenreported in the literature [reviewed in: Poirier D. (2003) Curr Med.Chem. 10:453-77]. In addition, the international patent application WO02/26706 discloses 17β-HSD2 inhibitors of non-steroidal origin.

Some thienopyrimidinones derivatives that are described as being usefulin therapy have already been disclosed in the literature: The Germanpatent application DE 2411273 (Schering AG) discloses compounds havinganti-inflammatory activity. Manhas et al describe the synthesis andanti-inflammatory activity of some substituted thienopyrimidinones[Manhas M S et al. (1972) J Med. Chem. 15(1):106-7]. Kapustina et al.describe the synthesis and antibacterial and chemotherapeutic orantitubecular activity of some substituted thienopyrimidones [KapustinaM V et al. (1992) Khimiko-Farmatsevticheskii Zhurnal 26(1):56-7; andKapustina M V et al (1991) Khimiko-Farmatsevticheskii Zhurnal 25(7):38-9].

Furthermore, several other Thienopyrimidinones derivatives have beendescribed but were not related to any medical use so far. For example,the compounds1,2,7,8,9,10,11,13-octahydro-13-oxo-4-(phenylthio)-[1]benzothieno[2′,3′:4,5]pyrimido[1,2-a]azepine-3-carboxaldehyde(CAS reg. no. 333774-42-8) and1,2,7,8,9,10,11,13-octahydro-13-oxo-4-(chloro)-[1]benzothieno-[2′,3′:4,5]pyrimido[1,2-a]azepine-3-carboxaldehyde(CAS reg. no 299962-60-0) are commercially available. Furthersubstituted thienopyrimidones have already been disclosed, e.g.:

-   1,2,7,8,9,10,11,13-octahydro-4-hydroxy-[1]benzothieno[2′,3′:4,5]pyrimido[1,2-a]azepin-13(7H)-one    (CA reg. no. 333774-26-8);-   2,3,8,9,10,11-hexahydro-[1]benzothieno[2′,3′:4,5]pyrimido[1,2-a]azepine-4,13(1H,7H)-dione    (CA reg. no. 141581-80-8);-   2,3,8,9-Tetrahydro[1]benzothieno[2,3-d]pyrrolo[1,2-a]pyrimidine-6,10(1H,7H)-dione    (CA reg. no. 141581-81-9),-   8,9,10,11-tetrahydro-4-hydroxy-[1]benzothieno[2′,3′:4,5]pyrimido[1,2-a]azepin-13(7H)-one    (CA reg. no. 333780-19-1);-   3-Butyl-2,7-dimethyl-4b,5,6,7,8,8a-hexahydro-3H-benzo[4,5]thieno[2,3-d]pyrimidin-4-one    (CA reg. no. 39625-80-4);-   1,2,3,4,5,8,9,10,11,12-Decahydro-14H-cyclohepta[4′,5′]thieno[2′,3′:4,5]pyrimido-[1,2-a]azepin-14-one-4-oxime    (CA reg. no. 299962-59-7);-   1,2,3,4,5,8,9,10,11,12-Decahydro-14H-cyclohepta[4′,5′]thieno[2′,3′:4,5]pyrimido-[1,2-a]azepin-14-one-3-oxime    (CA reg. no. 296798-31-7);-   1,2,3,4,7,9,10,12-Octahydro-12-oxo-8H-[1]-benzothieno[2,3-d]pyrido[1,2-a]pyrimidine-7-carboxylic    acid ethyl ester (CA reg no. 329059-69-0);-   1,2,3,4-Tetrahydro-12H-[1]-benzothieno[2,3-d]pyrido[1,2-a]pyrimidin-12-one    (CA Reg. No. 60943-07-9), and-   3-Methyl-2,3,4,7,8,9,10,11-octahydro-[1]-benzothieno[2′,3′:4,5]pyrimido-[1,2-a]azepin-13(1H)-one    (CA Reg. No. 677320-14-8).

However, according to the inventors' knowledge none of the already knowncompounds described above has been described as useful in the treatmentand/or prevention of a steroid hormone dependent disease or disorder,particularly a steroid hormone dependent disease or disorder requiringthe inhibition of the 17β-hydroxysteroid dehydrogenase (17HSD) type 1,type 2 or type 3 enzyme.

There is a need for the development of compounds that are selectivelyinhibiting the 17β-HSD1, 17β-HSD3 and/or 17β-HSD2 enzyme, whiledesirably failing to inhibit substantially other members of the 17β-HSDprotein family, or other catalysts of sex steroid degradation oractivation. In particular, it is an aim of the present invention todevelop selective inhibitors of the 17β-HSD1 enzyme, whereby in additionthe compounds have no or only pure antagonistic binding affinities tothe estrogen receptor (both subtypes α and β).

SUMMARY OF THE INVENTION

Therefore, it is an object of the present invention to provide novelinhibitors of the 17β-HSD1 and 17β-HSD2 enzyme, which have valuablepharmacological properties and which are suited for the treatment ofestrogen dependent diseases and disorders. It is a further object of thepresent invention to develop novel inhibitors of the 17β-HSD3 enzyme,which have valuable pharmacological properties and which are suited forthe treatment of androgen dependent diseases and disorders.

It has now been found that the thiophenepyrimidinone derivatives of thepresent invention would be valuable in therapy, especially in thetreatment or prevention of steroid hormone dependent diseases ordisorders, such as steroid hormone dependent diseases or disordersrequiring the inhibition of 17β-hydroxysteroid dehydrogenase (HSD)enzymes. In particular, compounds of formula (I) represent potentinhibitors of the 17β-HSD1, 17β-HSD3 and/or 17β-HSD2 enzyme and possessvaluable pharmacological properties for the treatment and/or prophylaxisof malignant steroid dependent diseases or disorders such as breastcancer, prostate carcinoma, ovarian cancer, uterine cancer, endometrialcancer and endometrial hyperplasia, but also for the treatment and/orprophylaxis of benign steroid dependent diseases or disorders such asendometriosis, uterine fibroids, uterine leiomyoma, adenomyosis,dysmenorrhea, menorrhagia, metrorrhagia, prostadynia, benign prostatichyperplasia, prostatitis, acne, seborrhea, hirsutism, androgenicalopecia, precocious puberty, adrenal hyperplasia, polycystic ovariansyndrome, or urinary dysfunction. Further estrogen-dependent diseaseswhich may be treated and/or prevented with an effective amount of acompound of the invention are multiple sclerosis, rheumatoid arthritis,Alzheimer's disease, colon cancer, tissue wounds, skin wrinkles andcataracts. Furthermore, compounds of formula (I) may be useful for theprevention and treatment of osteoporosis, and for blockingspermatogenesis and as an anti-fertility agent for males.

Accordingly, the present invention relates to the use of a compoundhaving the structural formula (I)

wherein

-   R₁ and R₂ represent the same or different alkyl, or one is alkyl and    the other is H, or-   R₁ and R₂ form together with their binding sites a cyclic 5-, 6-, 7-    or 8-membered ring system,-   which is saturated or contains one or more double bonds between the    ring atoms, and-   which ring optionally contains up to two heteroatoms in addition to    the nitrogen atom where R₁ is attached, the number of N atoms being    0-2 and the number of O or S atoms each being 0-1,-   wherein said ring is optionally substituted with up to three    substituents independently selected from the group consisting of    alkyl, substituted alkyl, aryl, or arylalkyl, whereby the aryl group    is optionally substituted, alkoxy, aryloxy, acyloxy, arylthio,    alkylthio, arylsulfonyl, alkylsulfonyl, hydroxyl, oxo, halogen,    amino, oxime, acyl, carboxyl, thiocarboxyl, and amido;-   R₃ and R₄ form together with their binding sites a cyclic 5-, 6-, 7-    or 8-membered hydrocarbon ringsystem, which is saturated or contains    one or more double bonds between the carbon atoms, and-   wherein said ring is optionally substituted with up to three    substituents independently selected from the group consisting of    alkyl, substituted alkyl, aryl or arylalkyl, whereby the aryl group    is optionally substituted, alkoxy, aryloxy, acyloxy, arylthio,    alkylthio, arylsulfonyl, alkylsulfonyl, hydroxyl, oxo, halogen,    amino, oxime, acyl, carboxyl, thiocarboxyl, and amido;-   provided that said compound is not    1,2,7,8,9,10,11,13-octahydro-13-oxo-4-(phenylthio)-[1]benzothieno[2′,3′:4,5]pyrimido[1,2-a]azepine-3-carboxaldehyde;    for the manufacture of a medicament for the treatment and/or    prevention of a steroid hormone dependent disease or disorder,    preferably for a steroid hormone dependent disease or disorder    requiring the inhibition of a 17β-hydroxysteroid dehydrogenase    (17β-HSD) enzyme, most preferably requiring the inhibition of the    17β-HSD type 1, 17β-HSD type 2 or 17β-HSD type 3 enzyme.

According to another aspect, the invention concerns a compound offormula (II)

wherein

-   R₁ and R₂ represent the same or different C₁-C₈-alkyl, or one is    C₁-C₈-alkyl and the other is H, or-   R₁ and R₂ form together with their binding sites a cyclic 5-, 6-, 7-    or 8-membered ring system,-   which is saturated or contains one or more double bonds between the    ring atoms, and-   which ring optionally contains up to two N-atoms in addition to the    nitrogen atom where R₁ is attached,-   wherein said ring is optionally substituted with up to two    substituents independently selected from the group consisting of    alkyl, substituted alkyl, aryl, or arylalkyl, whereby the aryl group    is optionally substituted, alkoxy, aryloxy, acyloxy, arylthio,    alkylthio, arylsulfonyl, alkylsulfonyl, hydroxyl, oxo, halogen,    amino, oxime, acyl, carboxyl, thiocarboxyl, and amido;-   the hydrocarbon chain —C(R5)-C(R6)-(CH)_(n)— of the ring-system    adjacent to the thiophen-ring is saturated or contains one or more    double bonds between the carbon atoms;-   n is an integer from 1 to 4, and-   R5 and R6 are individually selected from the group consisting of    hydrogen, alkyl, substituted alkyl, aryl or arylalkyl, whereby the    aryl group is optionally substituted, alkoxy, aryloxy, acyloxy,    arylthio, alkylthio, arylsulfonyl, alkylsulfonyl, hydroxyl, oxo,    halogen, amino, oxime, acyl, carboxyl, thiocarboxyl, and amido.    under the proviso that-   in case n represents 1, 2 or 3, and R1 and R2 independently are    selected from hydrogen or C₁-C₄ alkyl or together form an    unsubstituted alkylen group of 3-5 methylen groups or a iminoalkylen    group of 2-4 methylen groups in the alkylen group, optionally    substituted at the N-atom, then at least    -   (i) R5 or R6 has to be different from hydrogen, C₁-C₄ alkyl or        alkylcarboxyl, or    -   (ii) the hydrocarbon chain —C(R5)-C(R6)-(CH)n- of the        ring-system adjacent to the thiophen-ring has to be unsaturated        or aromatic;-   in case n represents 2 and R1-R2 form an unsubstituted alkylen group    of 3-5 methylen groups, then R6 has to be different from bromo,    dibromo or phenylthio, if R5 represents a hydroxyl or oxo group; or-   in case n represents 2 and R1-R2 form an unsubstituted pentamethylen    group, then R5 has to be different from phenylthio, if R6 represents    carbonyl,    for use in therapy.

According to a third aspect, the invention concerns a novel compound offormula (II)

wherein

-   R₁ and R₂ represent the same or different C₁-C₈-alkyl, or one is    C₁-C₈-alkyl and the other is H, or-   R₁ and R₂ form together with their binding sites a cyclic 5-, 6-, 7-    or 8-membered ring system,-   which is saturated or contains one or more double bonds between the    ring atoms, and-   which ring optionally contains up to two N-atoms in addition to the    nitrogen atom where R₁ is attached,-   wherein said ring is optionally substituted with up to two    substituents independently selected from the group consisting of    alkyl, substituted alkyl, aryl, or arylalkyl, whereby the aryl group    is optionally substituted, alkoxy, aryloxy, acyloxy, arylthio,    alkylthio, arylsulfonyl, alkylsulfonyl, hydroxyl, oxo, halogen,    amino, oxime, acyl, carboxyl, thiocarboxyl, and amido;-   the hydrocarbon chain —C(R5)-C(R6)-(CH)_(n)— of the ring-system    adjacent to the thiophen-ring is saturated or contains one or more    double bonds between the carbon atoms;-   n is an integer from 1 to 4, and-   R5 and R6 are individually selected from the group consisting of    hydrogen, alkyl, substituted alkyl, aryl or arylalkyl, whereby the    aryl group is optionally substituted, alkoxy, aryloxy, acyloxy,    arylthio, alkylthio, arylsulfonyl, alkylsulfonyl, hydroxyl, oxo,    halogen, amino, oxime, acyl, carboxyl, thiocarboxyl, and amido.    under the proviso that-   in case n represents 1, 2 or 3, and R1 and R2 are independently    selected from hydrogen or C₁-C₄ alkyl or form together an    unsubstituted alkylen group of 3-5 methylen groups or a iminoalkylen    group with 2-4 methylen groups in the alkylen group, optionally    substituted at the N-atom, then at least    -   (i) R5 or R6 has to be different from hydrogen, C₁-C₄ alkyl,        alkylcarboxyl, or ═N—OH, or    -   (ii) the hydrocarbon chain —C(R5)-C(R6)-(CH)_(n)— of the        ring-system adjacent to the thiophen-ring has to be aromatic,        and at least R5 or R6 has to be different from hydrogen;-   in case n represents 2 and R1-R2 form an unsubstituted alkylen group    of 3 or 5 methylen groups, then R6 has to be different from    hydrogen, bromo, dibromo or phenylthio, if R5 represents a hydroxyl    or oxo group;-   in case n represents 2 and R1-R2 form an unsubstituted pentamethylen    group, then R5 has to be different from phenylthio or chloro, if R6    represents carbonyl;-   in case n represents 2, the hydrocarbon chain —C(R5)-C(R6)-(CH)n- of    the ring-system adjacent the thiophene-ring is saturated and R1-R2    together with their binding sites form an unsubstituted    pyridine-ring, then at least one of R5 or R6 has to be different    from hydrogen; or-   in case —C(R5)-C(R6)-(CH)_(n)— represents an unsubstituted    tetramethylen group, R1-R2 have to be different from a tetramethylen    group substituted with a carboxylethylester group.

According to a fourth aspect, the invention concerns a pharmaceuticalcomposition comprising as active agent a compound of formula (II) asdefined herein, for which no use in therapy earlier has been disclosed,and at least a pharmaceutically acceptable carrier.

According to a fifth aspect, the invention concerns the use of acompound of formula (I), more preferably of formula (II), as definedherein, for the treatment or prevention of a steroid hormone dependentdisease or disorder. Preferably, the steroid hormone dependent diseaseor disorder is a disease or disorder requiring the inhibition of a17β-hydroxysteroid dehydrogenase enzyme, preferably of the 17β-HSD type1, 17β-HSD type 2 or 17β-HSD type 3.

According to a sixth aspect the invention concerns a method for thepreparation of the novel compounds of formula (I) wherein

a) a compound of formula 2

-   -   or a ring-substituted or ring-modified analogue thereof is        oxidized, preferably by subjecting to PCC and celite, to give an        oxo-substituted compound of formula 3 or an analogue thereof,

-   b) the oxo-substituted compound obtained in step a) is optionally    further subjected to Vilsmeier reaction, preferably by POCl₃-DMF, to    give a carbonylsubstituted compound of formula 4 or an analogue    thereof,

-   c) the chlorosubstituent in the carbonylsubstituted compound    obtained in step b) is optionally further replaced by an alkylthio    or an arylthio group by subjecting to an appropriate thiol in the    presence of a base to give an arylthio- or alkylthiosubstituted    compound of formula 5 or an analogue thereof,

-   d) the arylthio- or alkylthiosubstituted compound obtained in    step c) is optionally further    -   i) reduced to a compound of formula 6,

-   -   ii) reacted with NH2OH to give a compound of formula 7,        or

-   e) the compound obtained in step b) is optionally further    -   i) reduced so as to replace the carbonyl group with        hydroxyalkyl, or    -   ii) subjected to an appropriate thiol in the presence of a base        and acetone, so as to replace the chloro substituent by a thiol        group and to replace the carbonyl group with an oxosubstituted        alkenyl.        or-   f) the compound obtained in step a) is optionally further subjected    to DMF acetal so as to introduce a dimethylaminomethylene    substituent in the ring next to the oxo substituent.

DETAILED DESCRIPTION Definitions

The following terms are used to describe various constituents of thechemical composition useful in this invention. The terms are defined asfollows:

As used herein, the terms “comprising” and “including” are used hereinin their open, non-limiting sense.

The word “compound” shall here be understood to cover any and allisomers (e.g., enantiomers, stereoisomers, diastereomers, rotomers, andtautomers), racemates or any mixture of isomers, prodrugs, and anypharmaceutically acceptable salt of said compound.

Where the plural form is used for compounds, salts, and the like, thisis taken to mean also a single compound, salt, or the like.

The term “substituted” means that the specified group or moiety bearsone or more substituents. Where any group may carry multiplesubstituents and a variety of possible substituents is provided, thesubstituents are independently selected and need not be the same. Theterm “unsubstituted” means that the specified group bears nosubstituents. The term “optionally substituted” means that the specifiedgroup is unsubstituted or substituted by one or more substituents.

Any asymmetric carbon atoms may be present in the (R)-, (S)- or(R,S)-configuration, preferably in the (R)- or (S)-configuration,whichever is most active. Substituents at a double bond or a ring may bepresent in cis-(.═Z—) or trans (=E-) form.

The compounds of the present invention may contain asymmetric centers onthe molecule, depending upon the nature of the various substituents. Incertain instances, asymmetry may also be present due to restrictedrotation about the central bond adjoining the two aromatic rings of thespecified compounds. It is intended that all isomers (includingenantiomers and diastereomers), either by nature of asymmetric centersor by restricted rotation as described above, as separated, pure orpartially purified isomers or racemic mixtures thereof, be includedwithin the ambit of the instant invention.

The term “halogen” refers to fluorine (F, Fluoro-), bromine (Br,Bromo-), chlorine (Cl, Chloro), and iodine (J, Iodo-) atoms. Preferredin the context of the present invention are Br, Cl and F.

The terms “dihalogen”, “trihalogen” and “perhalogen” refer to two, threeand four substituents, respectively, each individually selected from thegroup consisting of fluorine, bromine, chlorine, and iodine atoms.

The term “hydroxyl” refers to the group —OH.

The term “oxo” refers to the group ═O.

The term “thio” refers to the group ═S.

The term “thiol” refers to the group —SH.

The term “sulfonyl” refers to the group —S(O)₁₋₂—.

For the purpose of the present invention, the carbon content of varioushydrocarbon containing moieties is indicated by a prefix designating theminimum and maximum number of carbon atoms in the moiety, i.e., theprefix C_(i)-C_(j) defines the number of carbon atoms present from theinteger “i” to the integer “j” inclusive. Thus C₁-C₄-alkyl refers toalkyl of 1-4 carbon atoms, inclusive, or methyl, ethyl, propyl, butyland isomeric forms thereof.

The term “alkyl” stands for a hydrocarbon radical which may be linear,cyclic or branched, with single or multiple branching, whereby the alkylgroup comprises 1 to 12 carbon atoms. In one embodiment, the term“alkyl” stands for a linear or branched (with single or multiplebranching) alkyl chain of 1 to 8 carbon atoms, exemplified by the term(C₁-C₈)alkyl, more preferably of 1 to 4 carbon atoms exemplified by theterm (C₁-C₄)alkyl. The term (C₁-C₈)alkyl is further exemplified by suchgroups as methyl; ethyl; n-propyl; isopropyl; n-butyl; sec-butyl;isobutyl; tert-butyl; n-pentyl; isopentyl; neopentyl; tert-pentyl; 2- or3-methylpentyl; n-hexyl; isohexyl, and the like. The alkyl group may bepartially unsaturated, forming such groups as, for example, methylenyl,ethenyl, ethylenyl, propenyl (allyl), methyl-propenyl, butenyl,pentenyl, pentinyl, hexenyl, octadienyl, and the like. The term “alkyl”further comprises cycloalkyl groups, preferably cyclo(C₃-C₈)alkyl whichrefers to cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl,cyclooctyl and isomeric forms thereof such as methylcyclopropyl; 2- or3-methylcyclobutyl; 2-, or 3-methylcyclopentyl, and the like. Thecycloalkyl group may also be partly unsaturated, forming such groups as,for example, cyclohexenyl, cyclopentenyl, cyclooctadienyl, and the like.Furthermore, the term “alkyl” comprises a cycloalkyl-alkyl groupcomprising 4 to 12 carbon atoms, preferably“cyclo(C₃-C₈)alkyl-(C₁-C₄)alkyl” which refers to a alkyl group of 1 to 4carbon atoms as described above substituted with a cyclo(C₃-C₈)alkylgroup as described above, forming such groups as for examplecyclopropylmethyl, cyclohexylmethyl, cyclopentylethyl orcyclohexenylethyl.

The term “substituted alkyl” refers to alkyl as just described andsubstituted by up to five, more preferably by up to three, mostpreferably by one or two substituents independently selected from thegroup consisting of halogen, hydroxyl, thiol, nitro, nitrile, alkoxy,aryloxy, acyloxy, amino, imino, oxime, amido, acylamino, alkylthio,arylthio, acyl, carboxyl, sulfamoyl, sulfonamide, and alkylsulfonyl, asdefined herein. These groups may be attached to any carbon atom of thealkyl moiety. Substituted alkyl is preferably substituted with hydroxyl,C₁-C₄-alkoxy, C₁-C₈-alkylthio, arylthio, preferably phenylthio, analkylacyl group —CO—R″, a carboxyl group —(C═O)—OR″, an alkylamino group—NR″₂, an alkylimino group ═N—R″, or an alkyloxime group ═N—O—R″,wherein R″ represents hydrogen or C₁-C₄-alkyl. Preferably substitutedalkyl refers to substituted C₁-C₄-alkyl, preferably methyl, substitutedmethylen and substituted C₂-C₄-alkenyl.

The term “alkoxy” refers to a group —OR, where R may be alkyl,arylalkyl, or substituted arylalkyl as defined herein, wherein the alkylchain may be optionally further substituted as defined herein.Preferably, the term “alkoxy” refers to —O—(C₁-C₄)alkyl (or(C₁-C₄)alkoxy), with the (C₁-C₄)alkyl group as defined above, or to—O—(C₁-C₄)alkyl-phenyl, preferably benzoxy or phenethyloxy, optionallysubstituted in the aryl group with up to five independently selectedsubstituents, in particular hydroxyl, halogen, (C₁-C₄)-alkyl, or(C₁-C₄)-alkoxy; the number of said substituents being up to five forhalogen, and up to three for any combination of said other substituents.

The term “aryloxy” refers to a group —OAr, where Ar may be aryl, orsubstituted aryl, as defined herein. Preferably, Ar represents aryl asdefined herein, which is optionally substituted in the aryl group withup to five independently selected substituents, in particular hydroxyl,halogen, (C₁-C₄)-alkyl, or (C₁-C₄)-alkoxy; the number of saidsubstituents being up to five for halogen, and up to three for anycombination of said other substituents. Preferably, aryloxy refers tophenoxy, optionally substituted as defined above.

The term “acyloxy” refers to a group —O—CO—R, where R may be alkyl,arylalkyl, substituted arylalkyl, aryl, or substituted aryl, as definedherein, wherein the alkyl chain may be optionally further substituted asdefined herein.

The term “alkylacyloxy” represents a preferred selection of the term“acyloxy” and refers to the group —O—CO—C₁-C₁₂-alkyl, preferably to—O—CO—C₁-C₈-alkyl, and most preferably to —O—CO—C₁-C₄-alkyl.

The term “arylacyloxy” represents a preferred selection of the term“acyloxy” and refers to the group —O—CO—Ar, wherein Ar represents arylas defined herein, preferably phenyl, which is optionally substituted inthe aryl group with up to five independently selected substituents, inparticular hydroxyl, halogen, (C₁-C₄)-alkyl, or (C₁-C₄)-alkoxy; thenumber of said substituents being up to five for halogen, and up tothree for any combination of said other substituents.

The term “acyl” refers to a group —(C═O)—R′, where R may be hydrogen,alkyl, aryl or aryl-(C₁-C₄)-alkyl (both optionally substituted in thearyl group with independently selected substituents as defined herein),as defined herein. Preferably, the term “acyl” refers to a group—(C═O)—R′, where R′ represents hydrogen, (C₁-C₄)alkyl, phenyl, orphenyl-(C₁-C₄)alkyl, preferably benzyl.

The term “carbonyl” represents a preferred selection of the term “acyl”and refers to the group —CHO.

The term “alkylacyl” represents a preferred selection of the term “acyl”and refers to a group —(C═O)-alkyl, preferably —(C═O)—(C₁-C₄)alkyl.

The term “arylacyl” represents a preferred selection of the term “acyl”and refers to the group —CO—Ar, wherein Ar represents aryl as definedherein, preferably phenyl, which is optionally substituted in the arylgroup as defined herein.

The term “carboxyl” refers to a group —(C═O)—OR, where R may behydrogen, alkyl, substituted alkyl, aryl or aryl-(C₁-C₄)-alkyl (bothoptionally substituted in the aryl group with independently selectedsubstituents as defined herein), as defined herein. Preferably, the term“carboxyl” refers to a group —(C═O)—OR′, where R′ represents hydrogen,(C₁-C₄)alkyl, phenyl, or (C₁-C₄)alkyl-phenyl, preferably benzyl; wherebythe phenyl moiety may be optionally substituted with substituentsindependently selected from the group consisting of hydroxyl, halogen,(C₁-C₄)alkoxy, and (C₁-C₄)-alkyl, the number of said substituents beingup to five for halogen, and up to three for any combination of saidother substituents.

The term “alkylcarboxyl” represents a preferred selection of the term“carboxyl” and refers to a group —(C═O)—OR, where R is hydrogen or C₁-C₄alkyl.

The term “thio-carboxyl” refers to a group —(C═O)—SR, where R may behydrogen, alkyl, substituted alkyl, aryl or aryl-(C₁-C₄)-alkyl (bothoptionally substituted in the aryl group with independently selectedsubstituents as defined herein), as defined herein. Preferably, the term“thio-carboxyl” refers to a group —(C═S)—OR′, where R′ representshydrogen, (C₁-C₄)alkyl, phenyl, or (C₁-C₄)alkyl-phenyl, preferablybenzyl; whereby the phenyl moiety may be optionally substituted withsubstituents independently selected from the group consisting ofhydroxyl, halogen, (C₁-C₄)alkoxy, and (C₁-C₄)-alkyl, the number of saidsubstituents being up to five for halogen, and up to three for anycombination of said other substituents.

The terms “alkylthio” (“alkylsulfanyl”) and “alkylsulfonyl” refers to agroup —SR and —S(O)_(n=1-2)—R, respectively, where R may be alkyl,substituted alkyl, arylalkyl, or substituted arylalkyl, as definedherein. Preferably, the term “alkylthio” (“alkylsulfanyl”) refers to agroup —SR′ and the term “alkylsulfonyl” refers to a group—S(O)_(n=1-2)—R′, respectively, where R′ represents (C₁-C₄)alkyl, or(C₁-C₄)alkyl-phenyl, preferably benzyl; optionally substituted in thealkyl chain with up to three substituents as defined herein, preferablyhydroxyl, (C₁-C₄)-alkoxy or halogen.

The term “arylthio” (“arylsulfanyl”) and “arylsulfonyl” refers to agroup —S—Ar and —S(O)_(n=1-2)—Ar, respectively, where Ar representsaryl, or substituted aryl, as defined herein. Preferably Ar representsaryl, which is optionally substituted in the aryl group withindependently selected substituents as defined herein, in particularhydroxyl, halogen, (C₁-C₄)-alkyl, or (C₁-C₄)-alkoxy, the number of saidsubstituents being up to five for halogen, and up to three for anycombination of said other substituents. Preferably, arylthio refers tophenylsulfanyl, optionally substituted as defined above.

The term “amino” refers to the group —NRR′, where R and R′ mayindependently be hydrogen, alkyl (optionally substituted in the alkylchain with up to five independently selected substituents as definedherein, in particular hydroxyl, halogen or (C₁-C₄)-alkoxy), aryl oraryl-(C₁-C₄)-alkyl (both optionally substituted in the aryl group withup to five independently selected substituents as defined herein, inparticular hydroxyl, halogen, (C₁-C₄)-alkyl, or (C₁-C₄)-alkoxy, thenumber of said substituents being up to five for halogen, and up tothree for any combination of said other substituents), as definedherein.

The term “alkylamino” represents a preferred selection of the term“amino” and refers to the group —NRR′, where R and R′ may independentlybe hydrogen or (C₁-C₄)alkyl.

The term “Imino” refers to the group ═NR, where R may be hydrogen, alkyl(optionally substituted in the alkyl chain with up to five independentlyselected substituents as defined herein, in particular hydroxyl, halogenor (C₁-C₄)-alkoxy), aryl or aryl-(C₁-C₄)-alkyl (both optionallysubstituted in the aryl group with up to five independently selectedsubstituents as defined herein, in particular hydroxyl, halogen,(C₁-C₄)-alkyl, or (C₁-C₄)-alkoxy, the number of said substituents beingup to five for halogen, and up to three for any combination of saidother substituents), as defined herein.

The term “alkylimino” represents a preferred selection of the term“imino” and refers to the group ═NR, where R may be hydrogen or(C₁-C₄)alkyl.

The term “oxime” refers to the group ═N—OR, where R may be hydrogen,alkyl (optionally substituted in the alkyl chain with up to fiveindependently selected substituents as defined herein, in particularhydroxyl, halogen or (C₁-C₄)-alkoxy), aryl or aryl-(C₁-C₄)-alkyl (bothoptionally substituted in the aryl group with up to five independentlyselected substituents as defined herein, in particular hydroxyl,halogen, (C₁-C₄)-alkyl, or (C₁-C₄)-alkoxy, the number of saidsubstituents being up to five for halogen, and up to three for anycombination of said other substituents), as defined herein.

The term “alkyl-oxime” represents a preferred selection of the term“oxime” and refers to the group ═N—O—R, where R may be hydrogen or(C₁-C₄)alkyl.

The term “amido” refers to the group —(C═O)—NRR′, where R and R′ mayindependently be hydrogen, alkyl (optionally substituted in the alkylchain with up to five independently selected substituents as definedherein, in particular hydroxyl, halogen or (C₁-C₄)-alkoxy), aryl oraryl-(C₁-C₄)-alkyl (both optionally substituted in the aryl group withindependently selected substituents as defined herein, in particularhydroxyl, halogen, (C₁-C₄)-alkyl, or (C₁-C₄)-alkoxy, the number of saidsubstituents being up to five for halogen, and up to three for anycombination of said other substituents), as defined herein.

The term “alkylamido” represents a preferred selection of the term“amido” and refers to the group —(C═O)—NRR′, where R and R′ may beindependently selected from hydrogen or (C₁-C₄)alkyl.

The term “aryl” refers to an aromatic carbocyclic group comprising 6 to14, more preferably 6 to 10, carbon atoms and having at least onearomatic ring or multiple condensed rings in which at least one ring isaromatic. Preferably, aryl is phenyl, naphthyl, indanyl, indenyl,fluorenyl, 1,2,3,4-tetrahydro-naphthalen-1-yl or even biphenyl.

The aryl group may optionally be substituted by substituentsindependently selected from the group consisting of halogen, hydroxyl,(C₁-C₆)alkoxy, (C₁-C₆)alkyl, oxo, thiol, carboxyl, aryloxy orarylalkyloxy (both optionally substituted in the aryl moiety withindependently selected substituents as defined herein),(C₁-C₆)alkylthio, arylthio or arylalkylthio (both optionally substitutedin the aryl moiety with independently selected substituents as definedherein), amino, amido, acyl, and acylamino, as defined herein, thenumber of said substituents being up to five for halogen, and up tothree for any combination of said other substituents.

Substituted aryl is preferably substituted by substituents selected fromthe group consisting of (C₁-C₆)alkoxy, preferably methoxy, hydroxyl,(C₁-C₄)alkyl, halogen, the number of said substituents being up to fivefor halogen, and up to four, preferably up to three, for any combinationof said other substituents. Preferably substituted aryl is substitutedphenyl.

The term “arylalkyl” refers to an alkyl group substituted with up tothree independently selected aryl groups; preferably the term“arylalkyl” refers to “aryl-(C₁-C₄)-alkyl” or diaryl-(C₁-C₄)-alkyl,whereby the aryl is an aryl group as defined above. Arylalkyl ispreferably benzyl (—CH2-Phenyl) or phenethyl (—CH₂—CH₂-Phenyl).

The term “substituted arylalkyl” refers to an arylalkyl group as definedabove, wherein the aryl group is substituted as defined above.

The term “pro-drug” as used herein, represents derivatives of thecompounds of the invention that are drug precursors which, followingadministration to a patient, release the drug in vivo via a chemical orphysiological process. In particular, pro-drugs are derivatives of thecompounds of the invention in which functional groups carry additionalsubstituents which may be cleaved under physiological conditions in vivoand thereby releasing the active principle of the compound (e.g., apro-drug on being brought to a physiological pH or through an enzymeaction is converted to the desired drug form).

The term “pharmaceutically acceptable salts” refers to salt forms thatare pharmacologically acceptable and substantially non-toxic to thesubject being administered the compounds of the invention.Pharmaceutically acceptable salts of compounds of formula I includeconventional and stoichiometrical acid-addition salts or base-additionsalts formed from suitable non-toxic organic or inorganic acids orinorganic bases. Acid addition salts, for example, from compounds offormula I with a basic nitrogen atom are formed preferably with organicor inorganic acids. Suitable inorganic acids are, for example, halogenicacids such as hydrochloric acid, sulfuric acid, or phosphoric acid.Suitable organic acids are, for example, carboxylic, phosphonic, orsulfonic acids, for example acetic acid, propionic acid, glycolic acid,lactic acid, hydroxybutyric acid, malic acid, malenic acid, malonicacid, salicylic acid, fumaric acid, succinic acid, adipic acid, tartaricacid, citric acid, glutaric acid, 2- or 3-glycerophosphoric acid andother mineral and carboxylic acids well known to those skilled in theart. The salts are prepared by contacting the free base forms with asufficient amount of the desired acid to produce a salt in theconventional manner. Compounds containing acidic substituents may alsoform salts with inorganic or organic bases. Examples of suitable basesfor salt formation include, but are not limited to, inorganic bases suchas alkali or alkaline earth-metal (e.g., sodium, potassium, lithium,calcium, or magnesium) hydroxides, and those derived from ammoniumhydroxides (e.g., a quaternary ammonium hydroxide such astetramethylammonium hydroxide). Also contemplated are salts formed withpharmaceutical acceptable amines such as ammonia, alkyl amines,hydroxyalkylamines, N-methylglucamine, benzylamines, piperidines, andpyrrolidines and the like. Certain compounds will be acidic in nature,e.g. those compounds which possess a carboxyl or phenolic hydroxylgroup. Salts of phenols can be made by heating acidic compounds with anyof the above mentioned bases according to procedures well known to thoseskilled in the art.

As used herein, the term “composition” is intended to encompass aproduct comprising the specified ingredients in the specified amounts,as well as any product which results, directly or indirectly, fromcombination of the specified ingredients in the specified amounts.

The phrase “effective amount” as used herein, means an amount of acompound or composition which is sufficient enough to significantly andpositively modify the symptoms and/or conditions to be treated (e.g.,provide a positive clinical response). The effective amount of an activeingredient for use in a pharmaceutical composition will vary with theparticular condition being treated, the severity of the condition, theduration of the treatment, the nature of concurrent therapy, theparticular active ingredient(s) being employed, the particularpharmaceutically acceptable excipient(s)/carrier(s) utilized, and likefactors within the knowledge and expertise of the attending physician.

Preferred Embodiments

According to a preferred embodiment of the present invention, thecompound used for the for the manufacture of a medicament for thetreatment and/or prevention of a steroid hormone dependent disease ordisorder, preferably for a steroid hormone dependent disease or disorderrequiring the inhibition of a 17β-hydroxysteroid dehydrogenase (17β-HSD)enzyme, most preferably requiring the inhibition of the 17β-HSD type 1,17β-HSD type 2 or 17β-HSD type 3 enzyme, is defined as follows: thecompound has the formula (II)

wherein

-   R₁ and R₂ represent the same or different C₁-C₈-alkyl, or one is    C₁-C₈-alkyl and the other is H, or-   R₁ and R₂ form together with their binding sites a cyclic 5-, 6-, 7-    or 8-membered ring system,-   which is saturated or contains one or more double bonds between the    ring atoms, and-   which ring optionally contains up to two N-atoms in addition to the    nitrogen atom where R₁ is attached,-   wherein said ring is optionally substituted with up to two    substituents independently selected from the group consisting of    alkyl, substituted alkyl, aryl, or arylalkyl, whereby the aryl group    is optionally substituted, alkoxy, aryloxy, acyloxy, arylthio,    alkylthio, arylsulfonyl, alkylsulfonyl, hydroxyl, oxo, halogen,    amino, oxime, acyl, carboxyl, thiocarboxyl, and amido;-   the hydrocarbon chain —C(R5)-C(R6)-(CH)_(n)— of the ring-system    adjacent to the thiophen-ring is saturated or contains one or more    double bonds between the carbon atoms;-   n is an integer from 1 to 4, and-   R5 and R6 are individually selected from the group consisting of    hydrogen, alkyl, substituted alkyl, aryl or arylalkyl, whereby the    aryl group is optionally substituted, alkoxy, aryloxy, acyloxy,    arylthio, alkylthio, arylsulfonyl, alkylsulfonyl, hydroxyl, oxo,    halogen, amino, oxime, acyl, carboxyl, thiocarboxyl, and amido;-   provided that said compound is not    1,2,7,8,9,10,11,13-octahydro-13-oxo-4-(phenylthio)-[1]benzothieno[2′,3′:4,5]pyrimido[1,2-a]azepine-3-carboxaldehyde;

According to a preferable embodiment the compound of the formula (II) ischaracterized in that

-   the cyclic 5-, 6-, 7- or 8-membered ring system formed together by    R₁ and R₂ is optionally substituted with up to two substituents    independently selected from the group consisting of oxo, —CO—R,    —CO—O—R, —O—R, —C₁-C₄-alkyl, optionally substituted with —O—R, —S—R    or —N(R)₂;-   R5 and R6 are individually selected from the group consisting of    hydrogen, oxo, halogen, —O—R′, —S—R′, —SO—R′, —CO—R, —CO—O—R, or    —C₁-C₄-alkyl, —C₁-C₄-alkenyl or ═C₁-C₄-alkylen, optionally    substituted in the alkyl chain with —O—R, —S—R, —N(R)₂, —CO—R, or    ═N—O—R,    wherein R represents hydrogen or C₁-C₄-alkyl; and    wherein R′ represents hydrogen, C₁-C₈-alkyl, which can be linear,    cyclic or branched; aryl-C1-C4-alkyl, preferably benzyl; or aryl,    preferably phenyl.

Especially preferable compounds are those wherein R₁ and R₂ formtogether with their binding sites an optionally substituted cyclic 5-,6-, 7- or 8-membered ring system, which is saturated or contains one ormore double bonds between the ring atoms, and which ring optionallycontains up to two N-atoms in addition to the nitrogen atom where R₁ isattached.

Particularly preferable compounds of formula (II) are those wherein R₅is selected from the group consisting of hydrogen, oxo, halogen, —OH,—O—C₁-C₄-alkyl; —S—C₁-C₄-alkyl, —S—C₃-C₈-cycloalkyl, —S-phenyl,—SO-phenyl.

Further preferable compounds of formula (II) are those wherein R6 isselected from the group consisting of hydrogen, carbonyl, alkylcarboxyl,preferably —COON, —C₁-C₄-alkyl, —C₁-C₄-alkenyl or ═C₁-C₄-alkylen,optionally substituted in the alkyl chain with —O—R, —N(R)₂, —CO—R, or═N—O—R, wherein R represents hydrogen or C₁-C₄-alkyl.

In a preferred embodiment, the invention relates to a compound selectedfrom the group consisting of exemplary compounds

-   2,3,8,9,10,11-Hexahydro[1]Benzothieno[2′,3′:4,5]pyrimido[1,2-a]azepine-4,13(1H,7H)-dione;-   1,2,6,7,8,9,10-heptahydrocyclopenta[4′,5′]thieno[2′,3′:4,5]pyrimido[1,2-a]-azepin-3,12-dione;-   1,2,3,4,8,9,10,11,12-nonahydrocyclohepta[4′,5]thieno[2′,3′:4,5]pyrimido-[1,2-a]azepine-5(5aH),14-dione;-   1,2,7,8,9,10,11,12-octahydro[1]benzothieno[2′,3′:4,5]pyrimido-[1,2-a]azocin-4,14(3H)-dione;-   1,2,3,4,7,8,9,10-Octahydro-12H-[1]benzothieno[2,3-d]pyrido[1,2-a]pyrimidin-12-one;-   5,6-Dihydro-2,3-dimethyl[1]benzothieno[2,3-d]pyrimidin-4,8(3H,7H)-dione;-   5,6-Dihydro-3-methyl[1]benzothieno[2,3-d]pyrimidin-4,8(3H,7H)-dione;-   5,6-Dihydro-3-ethyl-2-methyl[1]benzothieno[2,3-d]pyrimidin-4,8(3H,7H)-dione;-   4-Chloro-1,2,3,7,8,9,10,11,12-octahydro[1]benzothieno[2′,3′:4,5]pyrimido-[1,2-a]-azocin-14-one-4-carboxyaldehyde;-   1,2,3,4,5,8,9,10,11,12-Decahydro-14H-cyclohepta[4′,5′]thieno[2′,3′:4,5]-pyrimido[1,2-a]azepin-14-one;-   8-Chloro-5,6-dihydro-3-methyl[1]benzothieno[2,3-d]pyrimidin-4(3H)-one-7-carboxyaldehyde;-   1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(ethylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-carboxaldehyde;-   1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(propylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-carboxaldehyde;-   1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(butylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-carboxaldehyde;-   1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(isopropylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-carboxaldehyde;-   1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(t-butylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-carboxaldehyde;-   1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(cyclopentylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-carboxaldehyde;-   1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(cyclohexylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-carboxaldehyde;-   1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(phenylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-hydroxymethyl;-   1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(cyclohexylthio)[1]benzothieno-[2′,3′:4,5]pyrimido    [1,2-a]azepine-3-hydroxymethyl;-   Octahydro-13-oxo-4-(phenylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-oxime-methyl;-   4-Chloro-1,2,7,8,9,10,11,13-octahydro-13-oxo[1]benzothieno[2′,3′:4,5]-pyrimido[1,2-a]azepine-3-hydroxymethyl;-   3-N,N-Dimethylamino-methylen-2,3,8,9,10,11-hexahydro[1]benzothieno[2′,3′:4,5]pyrimido[1,2-a]azepine-4,13(1H,7H)-dione;-   1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(propylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-(3-oxo)but-1-ene;    and-   1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(butylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-(3-oxo)but-1-ene,    or a physiologically acceptable salt thereof.

Pharmaceutically acceptable salts of the compounds of the invention aswell as commonly used pro-drugs and active metabolites of thesecompounds are also within the scope of the invention.

The invention also relates to pharmaceutical compositions comprising oneor more of the compounds of the invention for which no use in therapyhas been disclosed earlier, or their salts or pro-drugs, as active agentand at lease one pharmaceutically acceptable carrier.

Furthermore, the invention relates to the use of an effective amount ofa novel compound as defined herein for the treatment or prevention of asteroid hormone dependent disease or disorder in a mammal, in particulara human. Preferably the steroid hormone dependent disease or disorder isan estradiol or testosterone dependent disease or disorder.

In a preferred embodiment, the invention relates to the use of aneffective amount of a novel compound as defined within the presentinvention for the treatment or prevention of a steroid hormone dependentdisease or disorder in a mammal, whereby the steroid hormone dependentdisease or disorder requires the inhibition of a 17β-hydroxysteroiddehydrogenase (HSD) enzyme, preferably the human 17β-hydroxysteroiddehydrogenase (HSD) enzyme type 1, type 2 or type 3.

In a further preferred embodiment of the invention the steroid hormonedependent disease or disorder to be treated and/or prevented requiresthe lowering of the endogenous 17β-estradiol or testosteroneconcentration in a generalized and/or tissue specific manner.

The invention also relates to a method of treating a mammal such as ahuman having a condition related to 17β-hydroxysteroid dehydrogenase(HSD) type 1, type 2 or type 3 activity, comprising administering to themammal an amount of a compound of this invention, or a salt or a prodrugthereof, which amount is effective to treat the condition.Administration of compounds of this invention in combination with otherpharmaceuticals used in treatment of the listed conditions iscontemplated.

The conditions to be treated and/or prevented in the context of thepresent invention include but are not limited to breast cancer, prostatecarcinoma, ovarian cancer, uterine cancer, endometrial cancer,endometrial hyperplasia, endometriosis, uterine fibroids, uterineleiomyoma, adenomyosis, dysmenorrhea, menorrhagia, metrorrhagia,prostadynia, benign prostatic hyperplasia, prostatitis, acne, seborrhea,hirsutism, androgenic alopecia, precocious puberty, adrenal hyperplasia,polycystic ovarian syndrome, and urinary dysfunction. A furthercondition to be treated and/or prevented in the context of the presentinvention includes osteoporosis.

Further estrogen-dependent diseases which may be treated and/orprevented with an effective amount of a compound of the invention aremultiple sclerosis, rheumatoid arthritis, Alzheimer's disease, coloncancer, tissue wounds, skin wrinkles and cataracts.

In a preferred embodiment the invention relates to use of an effectiveamount of a compound of the invention for the treatment or prevention ofone of the aforementioned disease or disorders in a mammal whereby themammal is a human, preferably a female and most preferably a pre- orperi-menopausal female in the case of gynecological disorders.

Furthermore, compounds of formula (I) may be useful for blockingspermatogenesis and as an anti-fertility agent for males.

The disclosed compounds are also useful as diagnostic agents (e.g. indiagnostic kits or for use in clinical laboratories) for screening forthe presence or absence of 17β-hydroxysteroid dehydrogenase (HSD) type1, type 2 and/or type 3 activity.

It will be appreciated that the methods of the present invention can beincorporated in the form of a variety of embodiments, only a few ofwhich are disclosed herein. It will be apparent for the expert skilledin the field that other embodiments exist and do not depart from thespirit of the invention. Thus, the described embodiments areillustrative and should not be construed as restrictive.

Administration Forms

The method of the invention is primarily intended for treatment in amammal, preferably in humans and other primates, of steroid hormonedependent diseases or disorders, in particular estradiol dependentdiseases or disorders, wherein the steroid hormone dependent disease ordisorder preferably requires the inhibition of a 17β-hydroxysteroiddehydrogenase (HSD) enzyme, preferably the type 1 17β-hydroxysteroiddehydrogenase (HSD) enzyme [EC 1.1.1.62].

The compounds may be administered orally, dermally, parenterally, byinjection, by pulmonal or nasal delivery, or sublingually, rectally orvaginally in dosage unit formulations. The term “administered byinjection” includes intravenous, intraarticular, intramuscular (e.g. bydepot injection where the active compounds are released slowly into theblood from the depot and carried from there to the target organs),intraperitoneal, intradermal, subcutaneous, and intrathecal injections,as well as use of infusion techniques. Dermal administration may includetopical application or transdermal administration. One or more compoundsmay be present in association with one or more non-toxicpharmaceutically acceptable auxiliaries such as excipients, adjuvants(e.g. buffers), carriers, inert solid diluents, suspensing agents,preservatives, fillers, stabilizers, anti-oxidants, food additives,bioavailability enhancers, coating materials, granulating anddisintegrating agents, binding agents etc., and, if desired, otheractive ingredients.

The pharmaceutical composition may be formulated for example asimmediate release, sustained release, pulsatile release, two or morestep release, depot or other kind of release formulations.

The manufacture of the pharmaceutical compositions according to theinvention may be performed according to methods known in the art andwill be explained in further detail below. Commonly known and usedpharmaceutically acceptable auxiliaries as well as further suitablediluents, flavorings, sweetening agents, coloring agents etc. may beused, depending on the intended mode of administration as well asparticular characteristics of the active compound to be used, such assolubility, bioavailability etc. Suitable auxiliaries and furtheringredients may be such as recommended for pharmacy, cosmetics andrelated fields and which preferably are listed in the EuropeanPharmacopoeia, FDA approved or cited in the “GRAS” list (FDA List offood additives that are ‘generally recognized as safe’ (GRAS)).

One mode of application of the compounds of general formula (I) or ofpharmaceutical compositions comprising one or more of said compounds isoral application, e.g., by tablets, pills, dragees, hard and soft gelcapsules, granules, pellets, aqueous, lipid, oily or other solutions,emulsions such as oil-in-water emulsions, liposomes, aqueous or oilysuspensions, syrups, elixiers, solid emulsions, solid dispersions ordispersible powders. For the preparation of pharmaceutical compositionsfor oral administration, the compounds suitable for the purposes of thepresent invention as defined above can be admixed with commonly knownand used adjuvants and excipients such as for example, gum arabic,talcum, starch, sugars (such as, e.g., mannitose, methyl cellulose,lactose), gelatin, surface-active agents, magnesium stearate, aqueous ornon-aqueous solvents, paraffin derivatives, cross-linking agents,dispersants, emulsifiers, lubricants, conserving agents, flavoringagents (e.g., ethereal oils), solubility enhancers (e.g., benzylbenzoate or benzyl alcohol) or bioavailability enhancers (e.g.Gelucire™). In the pharmaceutical composition, the active ingredientsmay also be dispersed in a microparticle, e.g. a nanoparticulate,composition.

For parenteral administration, the active agents can be dissolved orsuspended in a physiologically acceptable diluent, such as, e.g., water,buffer, oils with or without solubilizers, surface-active agents,dispersants or emulsifiers. As oils for example and without limitation,olive oil, peanut oil, cottonseed oil, soybean oil, castor oil andsesame oil may be used. More generally spoken, for parenteraladministration the active agent can be in the form of an aqueous, lipid,oily or other kind of solution or suspension or even administered in theform of liposomes or nano-suspensions.

Transdermal application can be accomplished by suitable patches, asgenerally known in the art, specifically designed for the transdermaldelivery of active agents, optionally in the presence of specificpermeability enhancers. Furthermore, also emulsions, ointments, pastes,creams or gels may be used for transdermal delivery.

Another suitable mode of administration is via intravaginal devices(e.g. vaginal rings) or intrauterine systems (IUS) containing reservoirsfor controlled release of active agents over extended periods of time.For rectal or vaginal administration of the drug the compounds may alsobe administered in the form of suppositories. These compositions can beprepared by mixing the drug with a suitable non-irritating excipientwhich is solid at ordinary temperatures but liquid at the rectal orvaginal temperature and will therefore melt in the rectum or vagina torelease the drug.

Another mode of application is by implantation of a depot implantcomprising an inert carrier material, such as biologically degradablepolymers or synthetic silicones such as e.g. silicone rubber. Suchimplants are designed to release the active agent in a controlled mannerover an extended period of time (e.g., 3 to 5 years).

It will be appreciated by those skilled in the art that the particularmethod of administration will depend on a variety of factors, all ofwhich are considered routinely when administering therapeutics.

The actually required dosages of the agents of this invention for anygiven patient will depend upon a variety of factors, including, but notlimited to the activity of the specific compound employed, theparticular HSD type 1, type 2 or type 3 related condition being treated,the particular composition formulated, the mode of administration, timeand duration of administration, route of administration and theparticular site being treated, and furthermore the age of the patient,the body weight of the patient, the general health of the patient, thegender of the patient, the diet of the patient, rate of excretion, drugcombinations, and the severity of the condition undergoing therapy.

It will be further appreciated by one skilled in the art that theoptimal course of treatment, i.e., the mode of treatment and the dailynumber of doses of a compound of Formula I or a pharmaceuticallyacceptable salt thereof given for a defined number of days, can beascertained by those skilled in the art using conventional treatmenttests. Optimal dosages for a given set of conditions may be ascertainedby those skilled in the art using conventional dosage-determinationtests in view of the experimental data for a given compound. For oraladministration, an exemplary daily dose generally employed will be fromabout 0.01 μg/kg to about 100 mg/kg of total body weight, wherebycourses of treatment may be repeated at appropriate time intervals.Administration of prodrugs may be dosed at weight levels that arechemically equivalent to the weight levels of the fully activecompounds. The daily dosage for parenteral administration will generallybe from about 0.01 μg/kg to about 100 mg/kg of total body weight. Adaily rectal dosage regimen will generally be from about 0.01 μg/kg toabout 200 mg/kg of total body weight. A daily vaginal dosage regimenwill generally be from about 0.01 μg/kg to about 100 mg/kg of total bodyweight. The daily topical dosage regimen will generally be from about0.1 μg to about 100 mg administered between one to four times daily. Thetransdermal concentration will generally be that required to maintain adaily dose of from 0.01 μg/kg to 100 mg/kg of total body weight.

Abbreviations and Acronyms

As used herein, the following terms have the indicated meanings:

-   20βP 20β-hydroxyprogesterone-   A 4-androstene-3,17-one-   Ac Acetyl-   AcOH acetic acid-   HSD hydroxysteroid dehydrogenase-   DHT dehydrotestosterone-   DMF N,N-dimethylformamide-   E1 estron-   E2 estradiol-   ER estrogen receptor-   EtOAc ethyl acetate-   GnRH Gonadotropin Releasing Hormone-   GRAS generally recognized as safe-   MS mass spectrometry-   NAD(P)[H]nicotinamide-adenine-dinucleotide (phosphate) [reduced    NAD(P)]-   NMR nuclear magnetic resonance-   P progesterone-   PCC pyridinium chlorochromate-   T testosterone-   TBAB Tetrabutylammonium Bromide-   THF tetrahydrofuran-   ‘TOF Time-of-flight’

Experimental Section General Preparative Methods

The compounds of the present invention may be prepared by use of knownchemical reactions and procedures. Nevertheless, the following generalpreparative methods are presented to aid the reader in synthesizing the17-13-hydroxysteroid dehydrogenase inhibitors with specific detailsprovided below in the experimental section to illustrate workingexamples. All variable groups of these methods are as described in thegeneric description if they are not specifically defined below.

It is recognized that compounds of the invention with each claimedoptional functional group may not be prepared by each of thebelow-listed methods. Within the scope of each method, optionalsubstituents may appear on reagents or intermediates which may act asprotecting or otherwise non-participating groups. Utilizing methods wellknown to those skilled in the art, these groups are introduced and/orremoved during the course of the synthetic schemes which provide thecompounds of the present invention.

The compounds according to this invention can be prepared as shown inSchemes 2 to 6 described in the Experimental section. It is evident thatotherwise ring-substituted or -modified compounds as defined by formula(I) of the claims can be prepared analogously, e.g. by usingring-substituted or—modified analogues of the starting compound (2) inScheme 1.

The invention will be illustrated by the following non-restrictiveExperimental Section. In order to more fully illustrate the nature ofthe invention and the manner of practicing the same, the followingexamples are presented, but they should not be taken as limiting.

Example 1

TABLE 1 Individual compounds of compound classes 2-4 prepared compound NR¹ R² 2a 2 —(CH₂)₅— 2b 1 —(CH₂)₅— 2c 3 —(CH₂)₅— 2d 2 —(CH₂)₆— 2e 2—(CH₂)₄— 2f 2 —(CH₂)₃— 2g 2 —CH₃ —CH₃ 2h 2 —CH₃ H 2i 2 —CH₃ —CH₂CH₃ 3a 2—(CH₂)₅— 3b 1 —(CH₂)₅— 3c 3 —(CH₂)₅— 3d 2 —(CH₂)₆— 3e 2 —(CH₂)₄— 3f 2—(CH₂)₃— 3g 2 —CH₃ —CH₃ 3h 2 —CH₃ H 3i 2 —CH₃ —CH₂CH₃ 4a 2 —(CH₂)₅— 4b 2—(CH₂)₆— 4c 2 —(CH₂)₃— 4d 3 —(CH₂)₅— 4e 2 —CH₃ H

General procedure for the synthesis of 2a-i 2a2,3,4,7,8,9,10,11-Octahydro[1]benzothieno[2′,3′:4,5]pyrimido-[1,2-a]azepin-13(1H)-one

To ethyl 2-amino-4,5,6,7-tetrahydrobenzo[b]thiophene-3-carboxylate 1b(0.22 mol) and ε-caprolactam (0.33 mol) in dry dichloroethane (1500 ml),POCl₃ (0.27 mol) was added dropwise. The mixture was heated under refluxuntil no starting material was detected on TLC after which ⅘ of thesolvent was evaporated. Water (200 ml) was added and the solution wasmade basic with 20% KOH. The solution was extracted with CH₂Cl₂, washedwith brine and water and dried with Na₂SO₄. After filtration the solventwas evaporated. Recrystallisation from ethanol afforded 2a as whitecrystals in 90% yield.

¹H NMR δ 1.85 (10H, m), 2.75 (2H, m), 3.00 (4H, m), 4.35 (2H, m)

2b1,2,3,6,7,8,9,10-Octahydro-12H-cyclopenta[4′,5′]thieno[2′,3′:4,5]pyrimido[1,2-a]-azepin-12-one

2b was synthesized as described above but ethyl2-aminocyclopenta(b)-thiophene-3-carboxylate was used.

¹H NMR δ 1.83 (8H, m), 2.45 (2H, qn), 3.00 (4H, m), 4.37 (2H, m)

2c1,2,3,4,5,8,9,10,11,12-Decahydro-14H-cyclohepta[4′,5′]thieno[2′,3′:4,5]pyrimido-[1,2-a]azepin-14-one

2c was synthesized as described above but ethyl2-aminocyclo-hepta(b)thiophene-3-carboxylate was used.

¹H NMR δ 1.77 (12H, m), 2.82 (2H, m), 3.01 (2H, m), 3.35 (2H, m), 4.36(2H, m)

2d2,3,4,7,8,9,10,11,12-Decahydro[1]benzothieno[2′,3′:4,5]pyrimido-[1,2-a]azocin-14(1H)-one

2d was synthesized as described above but 2-azacyclooctanone was used.

¹H NMR δ 1.40 (2H, m), 1.56 (2H, m), 1.89 (8H, m), 2.76 (2H, m), 2.99(4H, m), 4.27 (2H, m)

2e1,2,3,4,7,8,9,10-Octahydro-12H-[1]benzothieno[2,3-d]pyrido[1,2-a]pyrimidin-12-one

2e was synthesized as described above but

-valerolactam was used.

¹H NMR δ 1.91 (8H, m), 2.75 (2H, m), 2.97 (4H, m), 4.02 (2H, t)

2f2,3,6,7,8,9-Hexahydro[1]benzothieno[2,3-d]pyrrolo[1,2-a]pyrimidin-10(1H)-one

2f was synthesized as described above but 2-pyrrolidinone was used.

¹H NMR δ 1.86 (4H, m), 2.28 (2H, qn), 2.76 (2H, m), 3.00 (2H, m), 3.14(2H, t), 4.16 (2H, t)

2g5,6,7,8-Tetrahydro-2,3-dimethyl[1]benzothieno[2,3-d]pyrimidin-4(3H)-one

2g was synthesized as described above but N-methylacetamide was used.

¹H NMR δ 1.86 (4H, m), 2.58 (3H, s), 2.76 (2H, m), 3.00 (2H, m), 3.56(3H, m)

2h 5,6,7,8-Tetrahydro-3-methyl[1]benzothieno[2,3-d]pyrimidin-4(3H)-one

2h was synthesized as described above but acetamide was used.

¹H NMR δ 1.87 (4H, m), 2.78 (2H, m), 3.03 (2H, m), 3.56 (3H, s), 7.91(1H, s)

2i5,6,7,8-Tetrahydro-2-ethyl-3-methyl[1]benzothieno[2,3-d]pyrimidin-4(3H)-one

2i was synthesized as described above but N-methylpropanamide was used.

¹H NMR δ 1.36 (3H, t), 1.85 (4H, m), 2.78 (4H, m), 3.01 (2H, m), 3.57(3H, s)

General Procedure for the Synthesis of 3a-i:

PCC (37 mmol), Celite (20 g) and compound 2a (7.3 mmol) were mix into afine powder and dry benzene (150 ml) was added. The reaction mixture washeated under refluxed over night. The cooled slurry was filtratedthrough a pad of Celite and the solvent was evaporated.Recrystallisation from methanol afforded white powder in 50% yield.

3a2,3,8,9,10,11-Hexahydro[1]Benzothieno[2′,3′:4,5]pyrimido[1,2-a]azepine-4,13(1H,7H)-dione

¹H NMR δ 1.58 (6H, m), 2.23 (2H, qn), 2.67 (2H, t), 3.07 (2H, m), 3.28(2H, t), 4.36 (2H, m)

3b1,2,6,7,8,9,10-heptahydrocyclopenta[4′,5′]thieno[2′,3′:4,5]pyrimido[1,2-a]-azepin-3,12-dione

¹H NMR δ 1.88 (4H, m), 2.92 (2H, m), 3.09 (2H, m), 3.33 (2H, m), 4.39(2H, m)

3c1,2,3,4,8,9,10,11,12-nonahydrocyclohepta[4′,5]thieno[2′,3′:4,5]pyrimido-[1,2-a]azepine-5(5aH),14-dione

¹H NMR δ 1.90 (12H, m), 2.83 (2H, m), 3.05 (2H, m), 3.57 (2H, m), 4.36(2H, m)

3d1,2,7,8,9,10,11,12-octahydro[1]benzothieno[2′,3′:4,5]pyrimido-[1,2-a]azocin-4,14(3H)-dione

¹H NMR δ 1.45 (2H, m), 1.61 (2H, m), 1.97 (4H, m), 2.24 (2H, qn), 2.67(2H, m), 3.02 (2H, m), 3.29 (2H, t), 4.31 (2H, m)

3e1,2,3,4,7,8,9,10-Octahydro-12H-[1]benzothieno[2,3-d]pyrido[1,2-a]pyrimidin-12-one

¹H NMR δ 1.99 (4H, m), 2.23 (2H, qt), 2.66 (2H, t), 3.01 (2H, t), 3.28(2H, t), 4.03 (2H, t)

3f2,3,8,9-Tetrahydro[1]benzothieno[2,3-d]pyrrolo[1,2-a]pyrimidine-6,10(1H,7H)-dione

¹H NMR δ 2.29 (4H, m), 2.67 (2H, t), 3.24 (4H, m), 4.19 (2H, t)

3g5,6-Dihydro-2,3-dimethyl[1]benzothieno[2,3-d]pyrimidin-4,8(3H,7H)-dione

¹H NMR δ 2.24 (2H, qn), 2.63 (3H, s), 2.66 (2H, m), 3.28 (2H, t), 3.59(3H, s)

3h 5,6-Dihydro-3-methyl[1]benzothieno[2,3-d]pyrimidin-4,8(3H,7H)-dione

¹H NMR δ 2.25 (2H, qn), 2.69 (2H, t), 3.31 (2H, t), 3.60 (3H, s), 8.08(1H, s)

3i5,6-Dihydro-3-ethyl-2-methyl[1]benzothieno[2,3-d]pyrimidin-4,8(3H,7H)-dione

¹H NMR δ 1.39 (3H, t), 2.23 (2H, qn), 2.67 (2H, m), 2.81 (2H, q), 3.29(2H, t), 3.59 (3H, s)

General Procedure for the Synthesis of 4a-4-e:

POCl₃ (135 mmol) was added dropwise into DMF (170 mmol) and CH₂Cl₂ (3ml) at 0° C. After 30 minutes 3a (17 mmol) in CH₂Cl₂ (15 ml) was addeddropwise. The reaction mixture was allowed to reach room temperature andstirred for 48 h. The reaction was quenched with satured NaOAc,extracted with CH₂Cl₂, washed with brine and water, and dried withNa₂SO₄. After filtration the solvent was evaporated. The product waspurified by flash chromatography using CH₂Cl₂-EtOAc 9:1 as an eluent.Recrystallisation from ethanol afforded 4a as yellow crystals in 75%yield.

4a4-Chloro-1,2,7,8,9,10,11,13-octahydro-13-oxo[1]benzothieno[2′,3′:4,5]pyrimido[1,2-a]azepine-3-carboxaldehyde

¹H NMR δ 1.86 (2H, m), 2.81 (2H, dt), 3.07 (2H, m), 3.28 (2H, dt), 4.37(2H, m), 10.21 (1H, s)

4b4-Chloro-1,2,3,7,8,9,10,11,12-octahydro[1]benzothieno[2′,3′:4,5]pyrimido-[1,2-a]-azocin-14-one-4-carboxyaldehyde

Purification by flash chromatography afforded compound 4b as a majorproduct and compound 11 as a minor product.

¹H NMR δ 1.44 (2H, m), 1.60 (2H, m), 1.93 (4H, m), 2.76 (2H, m), 3.02(2H, m), 3.28 (2H, m), 4.31 (2H, m), 10.21 (1H, s)

114-Chloro-1,2,7,9,10,11,12,14-octahydro-14-oxo-8H-[1]benzothieno[2′,3′:4,5]pyrimido-[1,2-a]-azocine-3,7-dicarbaldehyde

¹H NMR δ (CDCl₃) 1.31 (2H, m), 1.82 (5H, m), 2.43 (1H, m), 2.81 (2H, t)3.29 (2H, m), 3.75 (2H, m), 4.93 (1H, m), 9.92 (1H, s), 10.21 (1H, s)

4c8-Chloro-4-oxo-1,2,3,4,8,9-hexahydro[1]benzothieno[2,3-d]pyrrolo[1,2-a]pyrimidine-7-carbaldehyde

¹H NMR δ 2.33 (2H, m), 2.81 (2H, m), 3.25 (4H, m), 4.20 (2H, m), 10.20(1H, s)

4d1,2,3,4,5,8,9,10,11,12-Decahydro-14H-cyclohepta[4′,5′]thieno[2′,3′:4,5]-pyrimido[1,2-a]azepin-14-one

¹H NMR δ 2.82 (2H, m), 3.31 (2H, m), 3.60 (3H, s), 8.04 (1H, s), 10.22(1H, s)¹H NMR δ 2.82 (2H, m), 3.31 (2H, m), 3.60 (3H, s), 8.04 (1H, s),10.22 (1H, s)

4e8-Chloro-5,6-dihydro-3-methyl[1]benzothieno[2,3-d]pyrimidin-4(3H)-one-7-carboxyaldehyde

¹H NMR δ 2.81 (2H, m), 3.31 (2H, m), 3.59 (3H, s), 8.03 (1H, s), 10.22(1H, s)

TABLE 2 Individual compounds of compound class 5 prepared. compound n R¹R² R⁷ 5a 2 —(CH₂)₅— -Ph 5b 2 —(CH₂)₅— —CH₂CH₃ 5c 2 —(CH₂)₅— —CH₂CH₂CH₃5d 2 —(CH₂)₅— —CH₂CH₂CH₂CH₃ 5e 2 —(CH₂)₅— —CH(CH₃)₂ 5f 2 —(CH₂)₅——C(CH₃)₃ 5g 2 —(CH₂)₅— cyclopentyl 5h 2 —(CH₂)₅— cyclohexyl 5i 2—(CH₂)₆— —CH₂CH₂CH₃General Procedure for the Synthesis of 5a-5h: 4a

(0.62 mmol) in THF (20 ml) followed by 1M NaOH (0.93 ml) were addeddropwise into thiophenol (1.06 mmol) in THF (2 ml) at −18° C. Thereaction mixture was allowed to reach room temperature and stirred untilno starting material was detected on TLC. Reaction mixture was pouredinto large excess of water and stirred for 1 h. The product wasfiltrated and recrystallisation afforded 5a as yellow crystals in 85%yield.

5a1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(phenylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-carboxaldehyde

¹H NMR δ 1.77 (6H, m), 2.82 (2H, m), 2.95 (2H, m), 3.25 (2H, m), 4.29(2H, m), 7.23 (5H, m), 10.47 (1H, s)

5b1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(ethylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-carboxaldehyde

5b was synthesized as described above but ethanethiol was used.

¹H NMR δ 1.26 (3H, t), 1.85 (6H, m), 2.76 (2H, m), 2.91 (2H, q), 3.07(2H, m), 3.22 (2H, m), 4.37 (2H, m), 10.48 (1H, s). M/z 360

5c1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(propylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-carboxaldehyde

5c was synthesized as described above but propanethiol was used.

¹H NMR δ 0.99 (3H, t), 1.63 (2H, m), 1.86 (6H, m), 2.76 (2H, m), 2.87(2H, t), 3.06 (2H, m), 3.22 (2H, m), 4.37 (2H, m), 10.49 (1H, s). M/z374

5d1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(butylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-carboxaldehyde

5d was synthesized as described above but butanethiol was used.

¹H NMR δ 0.89 (3H, t), 1.46 (4H, m), 1.86 (6H, m), 2.76 (2H, m), 2.89(2H, t), 3.06 (2H, m), 3.22 (2H, m), 4.37 (2H, m), 10.48 (1H, s). M/z388

5e1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(isopropylthio)[1]benzothieno-[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-carboxaldehyde

5e was synthesized as described above but 2-propanethiol was used.

¹H NMR δ 1.31 (6H, d), 1.85 (6H, m), 2.77 (2H, m), 3.06 (2H, m), 3.23(2H, m) 3.41 (1H, septet), 4.37 (2H, m), 10.47 (1H, m). M/z 374

5f1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(t-butylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-carboxaldehyde

¹H NMR δ 1.37 (9H, s), 1.86 (6H, m), 2.80 (2H, t), 3.06 (2H, m), 4.37(2H, m), 10.42 (1H, s)

5g1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(cyclopentylthio)[1]benzothieno-[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-carboxaldehyde

¹H NMR δ 1.60-1.85 (14H, m), 2.76 (2H, t), 3.22 (2H, t), 3.61 (1H, m),4.37 (2H, m), 10.46 (1H, s)

5h1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(cyclohexylthio)[1]benzothieno-[2′,′:4,5]-pyrimido-[1,2-a]azepine-3-carboxaldehyde

¹H NMR δ¹H NMR δ 1.25-1.99 (16H, m), 2.77 (2H, m), 3.06-3.27 (5H, m),4.36 (2H, m), 10.47 (1H, s)

5i1,2,7,9,10,11,12,14-octahydro-14-oxo-4-(propylthio)-8H-[1]benzothieno-[2′,3′:4,5]pyrimido-[1,2-a]-azocine-3-carbaldehyde

Compound 5i was synthesized as compound 5c using 4b as a startingmaterial. ¹H NMR (CDCl₃) δ0.99 (3H, t), 1.44 (2H, m), 1.62 (4H, m), 1.93(4H, m), 2.76 (2H, m), 2.87 (2H, m), 3.02 (2H, m), 3.22 (2H, m), 4.30(2H, broad s), 10.49 (1H, s)

Example 2

TABLE 3 Compound n R¹ R² R⁷ 6a 2 —(CH₂)₅— -Ph 6b 2 —(CH₂)₅— cyclohexyl6c 2 —(CH₂)₅— propyl 7 2 —(CH₂)₅— -PhGeneral Procedure for the Synthesis of 6a-6b and 8

NaBH₄ (0.66 mmol) was added into 5a (0.52 mmol) in ethanol (300 ml).After stirring for 20 minutes water (65 ml) was added followed byacidification with HCl. Ethanol was evaporated and the product wasfiltered affording 6a as pale yellow powder in 98% yield.

6a1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(phenylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-hydroxymethyl

¹H NMR δ 1.80 (6H, m), 2.79 (2H, m), 2.99 (2H, m), 3.29 (2H, m), 4.33(2H, m), 4.61 (2H, s), 7.17 (5H, m). M/z 410

6b1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(cyclohexylthio)[1]benzothieno-[2′,3′:4,5]pyrimido[1,2-a]azepine-3-hydroxymethyl

¹H NMR δ1.22-1.92 (16H, m), 2.66 (2H, m), 2.9-3.1 (3H, m), 3.21 (2H, m),4.36 (2H, m) 4.59 (2H, s)

6c1,2,8,9,10,11-Hexahydro-3-(hydroxymethyl)-4-(propylthio)-[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-13(7H)-one

Compound 6c was synthesized by the method described for 6a-b usingcompound 5c as a starting material.

¹H NMR (D₆-acetone) δ 0.96 (3H, t), 1.60 (2H, m), 1.84 (6H, m), 2.68(4H, m), 3.10 (4H, m), 4.38 (2H, m), 4.57 (2H, broad s)

MS (m/z) 376

Synthesis of 7

NaOAc (0.50 mmol) was added to NH₂OH.HCl (0.50 mmol) in absolute ethanol(3 ml) at 0° C., followed by addition of 5a (0.26 mmol) in dry THF (7ml). The reaction mixture was allowed to reach room temperature andstirred for over night. The reaction was quenched with water, extractedwith CH₂Cl₂, washed with brine and water, and dried with Na₂SO₄. Afterfiltration solvent was evaporated giving 7 in 95% yield.Recrystallisation from ethanol afforded pale yellow powder.

7Octahydro-13-oxo-4-(phenylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-oxime-methyl

¹H NMR δ 1.81 (6H, m), 2.96 (4H, m), 3.30 (2H, m), 4.34 (2H, m), 7.18(5H, m), 7.61 (1H, broad s), 8.76 (1H, s). M/z 423

Example 3

84-Chloro-1,2,7,8,9,10,11,13-octahydro-13-oxo[1]benzothieno[2′,3′:4,5]-pyrimido[1,2-a]azepine-3-hydroxymethyl

¹H NMR δ 1.84 (6H, m), 2.71 (2H, m), 3.05 (2H, m), 3.25 (2H, m), 4.36(2H, m), 4.45 (2H, s). M/z 366

Example 4

Scheme 4. Synthesis of Compound 9 Synthesis of 9

Dry DMF (5 ml), DMF acetal (1.56 mmol) and 3b (0.36 mmol) were refluxedfor 2 hours under a CaCl₂-tube, after which DMF was distilled away andthe crude product dried in a vacuum. Purification in a flash columnusing acetone as an eluent afforded 9 in 55% yield.

¹H NMR (CDCl₃) δ 1.86 (6H, m), 3.08 (2H, m), 3.17 (6H, s), 4.08 (2H, s),4.40 (2H, m), 7.41 (1H, s).

Example 5

General Procedure for the Synthesis of 10a-10b:

KOH (0.58 mmol) and EtSH (0.59 mmol) in EtOH (5 ml) were stirred for 30minutes, followed by addition of 4a in acetone (40 ml). After 30 minuteswater (250 ml) was added and the mixture stirred until the productprecipitates. Filtration afforded 10a as yellow powder in 86% yield.

10a1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(propylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-(3-oxo)but-1-ene

¹H NMR (CDCl₃) δ 1.00 (3H, t), 1.60 (2H, m), 1.85 (6H, m), 2.39 (3H, s),2.75 (4H, m), 3.05 (2H, m), 3.27 (2H, m), 4.37 (2H, m), 6.35 (1H, d),8.28 (1H, d)

10b1,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(butylthio)[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-(3-oxo)but-1-ene

¹H NMR (CDCl₃) δ 0.88 (3H, t), 1.46 (4H, m), 1.85 (6H, m), 2.38 (3H, s),2.76 (4H, m), 3.05 (2H, m), 3.27 (2H, m), 4.37 (2H, m), 6.35 (1H, d),8.27 (1H, d)

FURTHER EXAMPLES 127,8,9,10,11,13-Hexahydro-13-oxo-4-(phenylthio)-[1]benzothieno[2′,3′:4,5]-pyrimido[1,2-a]azepine-3-carbaldehyde

Compound 5a (100 mg, 245 mmol) and DDQ (67 mg, 295 mmol) were refluxedover night in 15 ml of dry benzene. The reaction mixture was cooled toroom temperature and filtered through a short column of silica gel. Thesolvent was evaporated. The product 12 was recrystallized fromEtOH/petrol ether. ¹H NMR (CDCl₃) δ 1.86 (6H, m), 3.12 (2H, m), 4.47(2H, m), 7.16 (5H, m), 8.16 (1H, d), 8.72 (1H, d), 10.78 (1H, s)

131,2,3,4,7,8,9,10,11,12-Decahydro-4-hydroxy-14H-[1]-benzothieno[2′,3′:4,5]-pyrimido[1,2-a]azocin-14-one

To compound 3d (200 mg, 0.66 mmol) in 5 ml of THF and 15 ml of EtOH,NaBH₄ (33 mg, 0.86 mmol) was added and the reaction mixture was stirredfor 15 minutes. The reaction was quenched with water and extracted withCH₂Cl₂. The organic layer was washed with brine and dried withNa_(a)SO₂. After filtration the solvent was evaporated. The product waspurified by flash chromatography using CH₂Cl₂/EtOAc 1:1 as an eluent.

¹H NMR (CDCl₃) δ 1.42 (2H, m), 1.59 (2H, m), 1.95 (7H, m), 2.14 (2H, m),3.05 (4H, m), 4.29 (2H, m), 4.87 (1H, m)

149-Methyl-2,3,4,7,8,9,10,11-octahydro-[1]benzothieno[2′,3′:4,5]pyrimido[1,2-a]azepin-13(1H)-one

The compound was synthesized by the method described for 2a-i using4-methyl-caprolactam as a starting material.

¹H NMR δ 0.98 (3H, d), 1.26 (2H, m), 1.84 (5H, m), 2.04 (2H, m), 2.74(2H, m), 3.01 (4H, m), 3.51 (1H, m), 5.17 (1H, m)

MS (m/z) 288

151,2,3,4,7,9,10,12-Octahydro-12-oxo-8H-[1]-benzothieno[2,3-d]pyrido[1,2-a]pyrimidine-7-carboxylicacid ethyl ester

The compound was synthesized by the method described for 2a-i using3-ethoxy-carbonyl-2-piperidone as a starting material.

¹H NMR δ 1.24 (3H, t), 1.99 (7H, m), 2.27 (1H, m), 2.74 (2H, m), 2.99(2H, m), 4.01 (3H, m), 4.20 (2H, q)

MS (m/z) 332

161,2,3,4,7,8,9,10,11,13-Decahydro-13-oxo-[1]benzothieno[2′,3′:4,5]pyrimido[1,2-a]azepine-9-carboxylicacid ethyl ester

The compound was synthesized by the method described for 2a-i usingethyl 7-oxo-4-azepanecarboxylate as a starting material.

¹H NMR δ 1.23 (3H, t), 1.88 (6H, m), 2.15 (2H, m), 2.72 (3H, m), 3.03(4H, m), 3.91 (1H, m), 4.13 (2H, q), 4.84 (1H, m)

172,3,8,9,10,11-Hexahydro-9-methyl-[1]-benzothieno[2′,3′:4,5]pyrimido[1,2-a]azepine-4,13(1H,7H)-dione

To compound 14 (4.0 g, 13.9 mmol), potassium peroxodisulfate (11.3 g,41.6 mmol) and CuSO₄.5H₂O, 250 ml of acetonitrile/water 1:1 were added.The reaction mixture was heated to reflux for 30 minutes. The reactionwas quenched with water and extracted with CH₂Cl₂. The organic layer waswashed with 10% sodium thiosulfate and brine, and dried over Na₂SO₄.After filtration the solvent was evaporated and the product purified byflash chromatography using CH₂Cl₂/EtOAc 9:1 as an eluent.

¹H NMR (CDCl₃) δ 1.00 (3H, d), 1.28 (2H, m), 1.90 (1H, m), 2.08 (2H, m),2.23 (2H, m), 2.66 (2H, m), 3.06 (2H, m), 3.28 (2H, m), 3.55 (1H, m),5.16 (1H, m)

MS (m/z) 302

184,12-Dioxo-1,2,3,4,7,9,10,12-octahydro-8H-[1]benzothieno[2,3-d]pyrido[1,2-a]pyrimidine-7-carboxylicAcid Ethyl Ester

Compound 18 was synthesized by the method described for compound 17using compound 15 as a starting material.

¹H NMR (CDCl₃) δ 1.26 (3H, t), 1.80 (1H, m), 2.18 (4H, m), 2.62 (3H, m),3.24 (3H, m), 3.81 (1H, m), 4.28 (3H, m)

194,13-Dioxo-1,2,3,4,7,8,9,10,11,13-decahydro-[1]benzothieno[2′,3′:4,5]-pyrimido[1,2-a]azepine-9-carboxylicAcid Ethyl Ester

Compound 19 was synthesized by the method described for compound 17using compound 16 as a starting material.

¹H NMR (CDCl₃) δ 1.28 (3H, t), 1.97 (2H, m), 2.21 (4H, m), 2.66 (2H, m),2.79 (1H, m), 3.03 (1H, m), 3.24 (3H, m), 4.01 (1H, m), 4.18 (2H, q),4.84 (1H, m)

204-Chloro-9-methyl-1,2,7,8,9,10,11,13-octahydro-13-oxo-[1]benzothieno-[2′,3′:4,5]pyrimido[1,2-a]azepine-3-carbaldehyde

POCl₃ (2.91 mmol) was added dropwise into DMF (3.02 mmol) in CHCl₃ (2ml) at 0° C. After 30 minutes compound 17 (0.36 mmol) in CHCl₃ (6 ml)was added slowly dropwise. The reaction mixture was allowed to reachroom temperature and stirred overnight after which the reaction mixturewas heated to 50° C. for 12 hours. The reaction mixture was cooled toroom temperature and quenched with saturated NaOAc, extracted withCH₂Cl₂, washed with brine and water, and dried over Na₂SO₄. Afterfiltration the solvent was evaporated. The product was purified by flashchromatography using CH₂Cl₂-EtOAc 9:1 as an eluent.

¹H NMR (CDCl₃) δ 1.00 (3H, d), 1.29 (2H, m), 1.90 (1H, m), 2.09 (2H, m),2.80 (2H, m), 3.06 (2H, m), 3.27 (2H, m), 3.55 (1H, m), 5.15 (1H, m),10.20 (1H, s)

214-Chloro-3-formyl-1,2,7,8,9,10,11,13-octahydro-13-oxo-[1]benzothieno-[2′,3′:4,5]pyrimido[1,2-a]azepine-9-carboxylicacid ethyl ester

Compound 21 was synthesized by the method described for 4a usingcompound 19 as a starting material, with the exception that the reactionmixture was stirred for 8 days.

¹H NMR (CDCl₃) δ 1.28 (3H, t) 1.98 (2H, m), 2.22 (2H, m), 2.79 (3H, m),3.03 (1H, m), 3.24 (3H, m), 4.01 (1H, m), 4.18 (2H, q), 4.85 (1H, m),10.20 (1H, s)

223-Formyl-1,2,7,8,9,10,11,13-octahydro-13-oxo-4-(propylthio)-[1]benzothieno[2′,3′:4,5]pyrimido[1,2-a]azepine-9-carboxylic acid ethyl ester

Compound 22 was synthesized by the method described for 5c usingcompound 21 as a starting material.

¹H NMR (CDCl₃) δ 0.99 (3H, t), 1.27 (3H, t), 1.62 (2H, m), 1.97 (2H, m),2.22 (2H, m), 2.77 (3H, m), 2.86 (2H, t), 3.02 (1H, m), 3.21 (3H, m),4.00 (1H, m), 4.17 (2H, q), 4.86 (1H, m), 10.49 (1H, s)

233-Formyl-1,2,7,8,9,10,11,13-octahydro-13-oxo-4-(propylthio)-[1]benzothieno-[2′,3′:4,5]pyrimido[1,2-a]azepine-9-carboxylic Acid Methyl Ester

Compound 2280.37 mmol) was dissolved into MeOH (10 ml), and KOH (0.98mmol) in MeOH (5 ml) was added. The reaction mixture was stirred for 2hours in room temperature. The reaction mixture was poured into water,acidified with HCl and extracted with CH₂Cl₂. Organic layer was washedwith brine and dried with Na₂SO₄. After filtration the solvent wasevaporated. Purification by flash chromatography using CH₂Cl₂-EtOAc 9:1as an eluent afforded compound 23 as a major product and compound 24 asa minor product. 23 ¹H NMR (CDCl₃) δ 0.99 (3H, t), 1.63 (2H, m), 1.98(2H, m), 2.23 (2H, m), 2.77 (3H, m), 2.86 (2H, t), 3.02 (1H, m), 3.21(3H, m), 3.73 (3H, s), 4.01 (1H, m), 4.84 (1H, m), 10.49 (1H, s)

243-Formyl-4-methoxy-1,2,7,8,9,10,11,13-octahydro-13-oxo-[1]benzothieno-[2′,3′:4,5]pyrimido[1,2-a]azepine-9-carboxylic Acid Methyl Ester

¹H NMR (CDCl₃) 1.98 (2H, m), 2.20 (2H, m), 2.72 (2H, m), 2.80 (1H, m),3.02 (1H, m), 3.21 (3H, m), 3.73 (3H, s), 4.04 (3H, s), 4.04 (1H, m),4.84 (1H, m), 10.19 (1H, s)

251,2,3,4-Tetrahydro-12H-[1]-benzothieno[2,3-d]pyrido[1,2-a]pyrimidin-12-one

2-Bromopyridine and ethyl2-amino-4,5,6,7-tetrahydrobenzo[b]thiophene-3-carboxylate 1b were heatedto 165° C. for three and half hours under argon. After cooling the solidmaterial was crystallized from EtOH. The crystallized product waspurified by flash chromatography using CH₂Cl₂/EtOAc 9:1 as an eluent.

¹H NMR (CDCl₃) δ 1.92 (4H, m), 2.81 (2H, m), 3.14 (2H, m), 7.00 (1H, m),7.57 (2H, m), 9.04 (1H, m)

MS (m/z) 256

262,3-Dihydro-12H-[1]-benzothieno[2,3-d]pyrido[1,2-a]pyrimidin-4,12(1H)-dione

Compound 26 was synthesized by the method described for compound 17using compound 25 as a starting material.

¹H NMR (CDCl₃) δ 2.30 (2H, m) 2.71 (2H, m), 3.40 (2H, t), 7.12 (1H, m),7.62 (1H, m), 7.76 (1H, m), 9.05 (1H, m)

273-Methyl-2,3,4,7,8,9,10,11-octahydro-[1]benzothieno[2′,3′:4,5]pyrimido-[1,2-a]azepin-13(1H)-one

Compound 27 was synthesized by the method described for 2a-i using2-amino-4,5,6,7-tetrahydro-6-methyl-benzo[b]thiophene-3-carboxylic acidethyl ester as a starting material.

¹H NMR (CDCl₃) δ 1.09 (3H, d), 1.44 (1H, m), 1.89 (8H, m), 2.37 (1H, m),2.85 (2H, m), 3.02 (2H, m), 3.21 (1H, m), 4.35 (2H, m)

MS (m/z) 288

282,3,8,9,10,11-Hexahydro-3-methyl-[1]-benzothieno[2′,3′:4,5]pyrimido[1,2-a]azepin-4,13(1H,7H)-dione

Compound 28 was synthesized by the method described for compound 17using compound 27 as a starting material.

¹H NMR (CDCl₃) δ 1.28 (3H, d), 1.85 (6H, m), 1.98 (1H, m), 2.29 (1H, m),2.67 (1H, m), 3.06 (2H, m), 3.13 (1H, m), 3.50 (1H, m), 4.36 (2H, m)

293-t-Butyl-2,3,4,7,8,9,10,11-octahydro-[1]-benzothieno[2′,3′:4,5]pyrimido-[1,2-a]azepin-13(1H)-one

Compound 29 was synthesized by the method described for 2a-i using2-amino-4,5,6,7-tetrahydro-6-t-butylbenzo[b]thiophene-3-carboxylic acidethyl ester as a starting material.

¹H NMR (CDCl₃) δ 0.95 (9H, s), 1.35 (1H, m), 1.55 (1H, m), 1.79 (6H, m),2.06 (1H, m), 2.51 (1H, m), 2.76 (2H, m), 3.01 (2H, broad s), 3.32 (1H,m), 4.35 (2H, m)

301,2,7,8,9,10,11,13-Octahydro-13-oxo-4-(phenylsulfinyl)-[1]benzothieno-[2′,3′:4,5]-pyrimido-[1,2-a]azepine-3-carbaldehyde

Compound 5a (100 mg, 0.25 mmol) and m-chloroperbenzoic acid (89 mg, 0.52mmol) in 25 ml dry CH₂Cl₂ were stirred for 3 days in room temperature.The reaction was quenched with water and extracted with CH₂Cl₂. Theorganic layer was washed with 10% sodium thiosulfate and brine and driedwith Na₂SO₄. After filtration the solvent was evaporated and the productpurified by flash chromatography using CH₂Cl₂/EtOAc 8:2

¹H NMR (CDCl₃) δ 1.80 (6H, m), 2.71 (2H, m), 3.03 (2H, m), 3.06 (1H, m),3.57 (1H, m), 4.31 (2H, m), 7.47 (3H, m), 7.69 (2H, m), 10.65 (1H, s)

314-Chloro-1,2,7,8,9,10,11,13-octahydro-13-oxo-[1]-benzothieno[2′,3′:4,5]-pyrimido[1,2-a]azepine-3-carboxylicAcid

Compound 4a (335 mg, 1.00 mmol) and 2-methyl-2-butene (1.06 ml, 10.01mmol) were dissolved into 50 ml THF. A freshly prepared solution of 80%NaClO₂ (339 mg, 3.00 mmol) and NaH₂PO₄.H₂O (414 mg, 3.00 mmol) in 55 mlof t-BuOH/H₂O 5:1 was added. The reaction mixture was stirred in roomtemperature for 6 hours. The reaction was quenched with water andextracted CH₂Cl₂. The organic phase was extracted with sat. NaHSO₄. Thewater phase was then made acidic with HCl and extracted with CH₂Cl₂. theorganic layer was washed with brine and dried over Na₂SO₂. Afterfiltration the solvent was evaporated ant the product recrystallizedfrom ethanol.

¹H NMR (D₆-DMSO) 1.71 (6H, m), 2.79 (2H, m), 3.06 (2H, m), 3.14 (2H, m),4.31 (2H, m), 13.07 (1H, broad s)

32 4-Hydroxy-3-methyl-2,3,4,7,8,9,10,11-octahydro[1]benzothieno[2′,3′:4,5]-pyrimido-[1,2-a]azepin-13(1H)-one

Compound 32 was synthesized as mixture of diastereomers by the methoddescribed for compound 13 using compound 28 as a starting material.

¹H NMR (CDCl₃) δ 1.14 (2H, d) 1.16 (2H, d), 1.58-2.04 (20H, m), 2.82(1H, m), 2.95 (1H, m), 3.04 (4H, m), 3.13 (1H, m), 3.25 (1H, m), 3.75(2H, m), 4.35 (4H, m), 4.42 (1H, m), 4.67 (1H, m)

333-Formyl-1,2,7,8,9,10,11,13-octahydro-13-oxo-4-(propylthio)-[1]benzothieno-[2′,3′:4,5]pyrimido[1,2-a]azepine-9-carboxylic Acid

Compound 22 (130 mg) in THF (2 ml) and 0.7 ml 10% KOH H₂O/MeOH (2:1)were stirred over night in room temperature. The reaction mixture waspoured into water and washed with ether. The water phase was acidifiedwith HCl and extracted with EtOAc. Organic phase was washed with brineand dried with Na₂SO₄. After filtration the solvent was evaporated andthe product recrystallized from ethanol.

¹H NMR (D₆-DMSO) δ 0.93 (2H, t), 1.55 (2H, m), 1.71 (2H, m), 2.14 (2H,m), 2.65 (2H, t), 2.67 (2H, m), 2.91 (2H, m), 3.10 (2H, m), 3.12 (2H,m), 3.97 (1H, m), 4.74 (1H, m), 10.36 (1H, s), 12.37 (1H, broad s); MS(m/z) 418

349-Methyl-1,2,7,8,9,10,11,13-octahydro-13-oxo-4-(propylthio)-[1]benzothieno-[2′,3′:4,5]-pyrimido[1,2-a]azepine-3-carbaldehyde

Compound 34 was synthesized by the method described for 5c usingcompound 21 as a starting material.

¹H NMR (CDCl₃) δ 0.99 (3H, t), 1.00 (3H, d), 1.28 (2H, m), 1.63 (2H, m),1.89 (1H, m), 2.08 (2H, m), 2.75 (2H, m), 2.86 (2H, t), 3.06 (2H, m),3.21 (2H, m), 3.55 (1H, m), 5.16 (1H, m), 10.49 (1H, s)

359-(Hydroxymethyl)-2,3,4,7,8,9,10,11-octahydro-[1]benzothieno[2′,3′:4,5]-pyrimido[1,2-a]azepin-13(1H)-one

Compound 16 (0.72 mmol) in dry THF (6 ml) was added dropwise to LiAlH₄(1.95 mmol) in THF (2 ml) under argon at 0° C. The reaction mixture wasstirred for 20 minutes. The reaction was quenched with water and 10%NaOH was added, and extracted with ether. The organic layer was washedwith brine and dried over Na₂SO₄. After filtration the solvent wasevaporated and the product purified by flash chromatography using EtOAcas an eluent.

¹H NMR (CDCl₃) δ 1.31 (2H, m), 1.80 (6H, m), 2.18 (2H, m), 2.75 (2H, m),3.01 (3H, m), 3.11 (1H, m), 3.51 (3H, m), 5.25 (1H, m)

It is evident that otherwise ring-substituted or -modified compounds asdefined by formula (I) of the claims can be prepared analogously, e.g.by using ring-substituted or -modified analogues of the startingcompound (2) in Scheme 2.

Further compounds of general formula IV falling under the scope ofgeneral formula I can prepared by parallel chemistry using a reaction asshown in the following scheme 6:

In a reaction vessel at room temperature are put together sequentially0.25 M lactam, 0.25 M amino ester and 0.25 M POCl₃. Of all reactants oneequivalent is used as solution or suspension in chlorobenzene. Aftershaking for 80 hours at 100° C., the mixtures are cooled to roomtemperature, washed with 5% NaOAc and extracted with EtOAc. The organiclayers are collected and concentrated to yield the desired compound. Theobtained material of the formula IV was thereafter analyzed by LC-MS.The LC-MS system consists of 2 Perkin Elmer series 200 micro-pumps. Thepumps are connected to each other by a 50 μl tee mixer. The mixer isconnected to the Gilson 215 auto-sampler. The LC methode consists of thefollowing steps:

Step total time flow (ul/min) A (%) B (%) 0 0 2300 95 5 1 1.8 2300 0 1002 2.5 2300 0 100 3 2.7 2300 95 5 4 3.0 2300 95 5 Solution A = 100% H₂Owith 0.025% HCOOH and 10 mmol NH₄HCOO pH = +/−3 Solution B = 100% MeOHwith 0.025% HCOOH

The auto sampler has a 2 μl injection loop. The auto sampler isconnected to a Varian Polaris C18 A 30*4.6 mm column with 3 μmparticles. The column is thermo stated in a Perkin Elmer series 200column oven at 40° C. The column is connected to an Applied BiosystemsABI 785 UV meter with a 2.7 μl flowcel. The wavelength is set to 254 nm.The UV meter is connected to a Sciex API 150EX mass spectrometer havingthe following parameters (Scanrange: 150-900 Amu, Polarity: positive,Scan mode: profile, Resolution Q1: UNIT, Step size: 0.10 amu, Time perscan: 0.500 sec, NEB: 10, CUR: 10, IS: 5200, TEM: 325, DF: 30, FP: 225,EP: 10). The light scattering detector is connected to the Sciex API150. The light scattering detector is a Sedere Sedex 55 operating at 50°C. and 3 bar N₂ pressure. The complete systems is controlled by a Delloptiplex GX400 computer operating under Windows NT.

The following table 4 lists compounds No. 41 to 53 of the generalformula IV, which were prepared according to Scheme 6 starting with thelactams and the amino acid esters. In addition, the Molecular Weight andthe Retention Time of the synthesized compounds determined by the LC-MSanalysis are shown.

TABLE 4 Compounds No. 41 to 53 of the general formula IV: No. StructuralFormula Amino ester Lactam MH+ RT 40

ETHYL 2-AMINO-4,5,6,7- TETRAHYDROBENZO[B] THIOPHENE-3- CARBOXLATE 2-AZACYCLOOCTANONE 288, 13 1,915 41

ETHYL 2-AMINO- BENZO[B]THIOPHENE-3- CARBOXYLATE 2- AZACYCLOOCTANONE 284,10 1,897 42

2-AMINO-7-HYDROXY- BENZO[B]THIOPHENE-3- CARBOXYLIC ACID ETHYL ESTER 2-AZACYCLOOCTANONE 300, 09 1,700 43

ETHYL 2-AMINO-7-OXO- 4,5,6,7-TETRAHYDRO-1- BENZOTHIOPHENE-3- CARBOXYLATE2- AZACYCLOOCTANONE 302, 11 1,684 44

ETHYL 2-AMINO- BENZO[B]THIOPHENE-3- CARBOXYLATE DELTA- VALEROLACTAM 256,07 1,736 45

ETHYL 2-AMINO- BENZO[B]THIOPHENE-3- CARBOXYLATE 3-CARBETHOXY-2-PIPERIDONE 328, 09 2,039 46

ETHYL 2-AMINO- BENZO[B]THIOPHENE-3- CARBOXYLATE EPSILON- CAPROLACTAM270, 08 1,858 47

2-AMINO-7-HYDROXY- BENZO[B]THIOPHENE-3- CARBOXYLIC ACID ETHYL ESTEREPSILON- CAPROLACTAM 286, 08 1,645 48

ETHYL 2-AMINO- BENZO[B]THIOPHENE-3- CARBOXYLATE N-ETHYL-2,3-DIKETOPIPERAZINE 299, 07 1,509 49

2-AMINO-7-HYDROXY- BENZO[B]THIOPHENE-3- CARBOXYLIC ACID ETHYL ESTERN-ETHYL-2,3- DIKETOPIPERAZINE 315, 07 1,386 50

ETHYL 2-AMINO-4,5,6,7- TETRAHYDROBENZO[B] THIOPHENE-3- CARBOXLATEN-ETHYL-2,3- DIKETOPIPERAZINE 303, 10 1,576 51

ETHYL 2-AMINO-7-OXO- 4,5,6,7-TETRAHYDRO-1- BENZOTHIOPHENE-3- CARBOXYLATEN-ETHYL-2,3- DIKETOPIPERAZINE 317, 08 1,277 52

ETHYL 2-AMINO- BENZO[B]THIOPHENE-3- CARBOXYLATE 4-METHOXY-3-PYRROLIN-2-ONE 270, 05 1,703 53

ETHYL 2-AMINO-4,5,6,7- TETRAHYDROBENZO[B] THIOPHENE-3- CARBOXLATE4-METHOXY-3- PYRROLIN-2-ONE 274,08 1,782

Biological Testing Materials and Methods

1. Inhibition of the 17β-hydroxysteroid Dehydrogenase Type 1, Type 2 andType 3 Enzyme

The compounds were screened in respect of 17β-HSD enzyme activity invitro on established MCF-7 cell lines, each stably expressing one of therespective 17β-HSD isoenzymes. The interconversion of substrate by eachisoenzyme and the 17β-HSD inhibiting activity of chemical compounds inthese cell lines were detected by HPLC system.

Varying amounts of the test compounds were incubated in the growthmedium of the 17β-HSD expressing cells together tritium labeledsubstrate (2 nM estrone for 17β-HSD type 1; 2 nM estradiol for 17β-HSDtype 2; and 2 nM androstenedione for 17β-HSD type 3). The medium sampleswere removed after exact incubation time and the reaction is stopped bytrichloroacetic acid (TCA). The samples were analyzed by HPLC-coupledflow scintillation analysis.

For each enzyme type, the HSD-inhibiting activity of an individual testcompound was calculated by comparing the conversion of a control samplewithout any test compound (referred to as “Negative Control”) to the(reduced) conversion of the test sample containing the particularcompound to be tested (referred to as “Test Sample”).

${\% \mspace{14mu} {inhibition}} = {100 \times \frac{\begin{matrix}{{{Conversion}\mspace{14mu} {in}\mspace{14mu} {Negative}\mspace{14mu} {Control}} -} \\{{Conversion}\mspace{14mu} {in}\mspace{14mu} {Test}\mspace{14mu} {Sample}}\end{matrix}}{{Conversion}\mspace{14mu} {in}\mspace{14mu} {Negative}\mspace{14mu} {Control}}}$

The results obtained are shown in the following Table 3. Twoconcentrations of each compound were used. The number of the compoundrefers to the numbers indicated in the Experimental Section.

TABLE 3 % Inhibition of the 17β-HSD enzymes type 1, type 2 and type 3 bythe compounds of the invention HSD1 HSD2 HSD3 Compound 1 μM 10 μM 1 μM10 μM 1 μM 10 μM No. 2a 10.0 45.9 16.2 23.4 39.0 24.9 No. 2b 21.0 15.015.9 21.4 4.2 9.5 No. 2c 22.7 39.8 8.9 13.5 50.8 47.0 No. 2e 15.8 20.026.0 28.9 46.8 38.8 No. 2f 19.9 19.2 37 32.4 48.2 40.4 No. 2g 30.7 37.044.9 24.9 51.7 51.3 No. 2i 12.7 18.5 20.6 29.4 4.4 9.9 No. 3b 13.5 20.714.1 25.4 0.3 13.7 No. 3d 14.3/18.2^(x)) 42.9/37.6^(x)) 14.1 11.115.3/22.3^(x))  16.7/8.1^(x)) No. 3e 6.3 35.8 25.7 16.3 38.5 47.4 No. 3f8.7 22.0 31.1 21.8 43.9 40.2 No. 4a 27.9 78.0 7.7 22.5 9.4 53.8 No. 4b36.9/31.2^(x)) 82.1/80.1^(x)) 3.5 1.5 5.3/14.3^(x)) 42.1/43.9^(x)) No.4e 23.5/24.6^(x)) 44.4/52.1^(x)) 9.6 28.3 9.8/11.1^(x)) 14.2/17.3^(x))No. 5b 66.1 100.0 19.7 25.2 43.4 100.0 No. 5c 74.0 100.0 8.9 18.6 10.5100.0 No. 5d 54.1 100.0 8.2 15.1 32.6 72.0 No. 5e 71.9 100.0 8.4 20.250.8 86.3 No. 5i 43.9 100.0 11.6 17.3 22.5 80.5 No. 6a 51.8 100.0 11.642.5 45.9 48.9 No. 6c 47.7 82.4 37.5 36.7 20.4 38.1 No. 11 4.7 51.1 16.728.3 7.3 31.7 No. 12 15.1 87.9 7.0 8.9 8.6 25.5 No. 13 5.0 20.0 1.7 18.6−2.2 11.7 No. 14 12.8 19.6 5.9 14.9 27.6 34.2 No. 15 14.8 24.5 10.7 15.7−0.3 13.5 No. 16 17.2 18.6 3.0 24.8 13.6 22.9 No. 17 18.4 39.4 15.2 20.121.6 32.4 No. 18 0.5 9.5 18.7 23.8 −3.0 21.3 No. 19 10.6 33.1 0.6 4.413.0 10.5 No. 20 10.4 45.7 11.2 6.2 32.0 100.0 No. 21 10.9 33.5 13.132.9 20.0 87.3 No. 22 24.3 46.5 6.8 16.1 25.5 100.0 No. 23 17.4 64.7 9.717.7 19.5 89.5 No. 24 24.9 32.3 3.1 19.3 18.0 59.6 No. 25 6.2 8.7 14.033.6 −3.2 7.9 No. 26 6.4 −0.8 15.1 16.4 −2.1 0.3 No. 27 11.4 25.6 5.33.2 22.9 24.9 No. 28 14.3 15.3 18.2 18.7 7.8 9.5 No. 29 −1.0 1.3 13.110.1 14.9 25.8 No. 30 23.1 49.0 12.8 24.8 11.1 31.2 No. 31 2.1 6.8 −0.29.3 −6.4 −3.9 No. 32 19.7 22.5 14.0 −5.1 n.d. n.d. No. 33 15.3 22.6 5.0−0.1 n.d. n.d. No. 34 14.9 48.1 13.0 −0.5 n.d. n.d. No. 35 4.8 22.4 15.827.7 16.2 23.4 ^(x))two tests were run

2. Estrogen Receptor Binding Assay

The binding affinity of the compounds of the invention to the estrogenreceptor α and to the estrogen receptor β may be determined according tothe in vitro ER binding assays described by Koffmann et al. [Koffmann Bet al. (1991) J. Steroid. Biochem. Mol. Biol. 38:135]. Alternatively, anestrogen receptor binding assay may be performed according tointernational patent application PCT/US/17799 (published as WO00/07996).

3. Estrogen Receptor Transactivation Assays

Compounds of the invention showing binding affinity towards the estrogenreceptor may be further tested with regard to their individualestrogenic or anti-estrogenic potential (agonistic binding orantagonistic binding to the ERα or ERβ). The determination of theestrogen receptor agonist activity may be performed according to an invitro assay system using the MMTV-ERE-LUC reporter system which is forexample described within U.S. patent application Ser. No. 10/289,079(published as US 2003/0170292):

To assay estrogen receptor agonist activity, Hela cells are grown in24-well microtiter plates and then transiently co-transfected with twoplasmids using lipofectamine. The first plasmid comprises DNA encodinghuman estrogen receptor (either ER-alpha or ER-beta), and the secondplasmid comprises an estrogen-driven reporter system comprising: aluciferase reporter gene (LUC) whose transcription is under the controlof upstream regulatory elements comprising 4 copies of the vitellogeninestrogen response element (ERE) cloned into the mouse mammary tumorvirus (MMTV) promoter (the full name for the reporter system being“MMTV-ERE-LUC”). Cells are exposed to the compounds of the invention inRPMI 1640 medium, supplemented with 10% charcoal-treated fetal calfserum, 2 mM L-glutamine, 0.1 mM non-essential amino acids and 1 mMsodium pyruvate for 42-48 hours at 37° C. in a 5% carbon dioxideincubator. Concurrently, cells exposed to estradiol (1 nM) serve aspositive controls. Replicate wells exposed to the solvent in which thecompounds of the invention are dissolved (i.e. ethanol or methanol) areused as negative controls. After the 42-48 hr incubation period, cellsare rinsed with phosphate buffered saline (PBS), lysis buffer (PromegaCorp) is added, and cell lysates are collected for measurement ofluciferase activity with a luminometer. Estrogenic activity of thecompounds of the invention is expressed as fold-increase in luciferaseactivity as compared to that observed in negative control cells.

Alternatively, the determination of the estrogen receptortransactivation activity (estrogenicity assay or agonist assay) and ofthe inhibitory potency of transactivation activity (anti-estrogenicityassay or antagonist assay) may be performed according to internationalpatent application PCT/US/17799 (published as WO 00/07996).

It will be appreciated that the methods of the present invention can beincorporated in the form of a variety of embodiments, only a few ofwhich are disclosed herein. It will be apparent for the expert skilledin the field that other embodiments exist and do not depart from thespirit of the invention. Thus, the described embodiments areillustrative and should not be construed as restrictive.

CITED LITERATURE

The following documents provide background information and are herebyincorporated herein by reference.

-   Labrie et al. (2000) “Role of 17 beta-hydroxysteroid dehydrogenases    in sex steroid formation in peripheral intracrine tissues” Trends    Endocrinol Metab., 11:421-7-   Labrie F et al. (1997) “The key role of 17 beta-hydroxysteroid    dehydrogenases in sex steroid biology.” Steroids, 62:148-58-   Tamaya et al. (1985) “Comparison of cellular levels of steroid    receptors in uterine leiomyoma and myometrium.” Acta Obstet Gynecol    Scand., 64:307-9-   Poirier D. (2003) “Inhibitors of 17 beta-hydroxysteroid    dehydrogenases” Curr Med. Chem. 10:453-77-   Geissler W M et al. (1994) “Male pseudohermaphroditism caused by    mutations of testicular 17beta-hydroxysteroid dehydrogenase 3.” Nat.    Genet., 7:34-9.-   Oefelein M G & Cornum R (2000) “Failure to achieve castrate levels    of testosterone during luteinizing hormone releasing hormone agonist    therapy: the case for monitoring serum testosterone and a treatment    decision algorithm.” J Urol.; 164:726-9.-   U.S. Pat. No. 6,541,463-   WO 01/42181-   WO 98/32724-   WO 98/30556-   WO 99/12540-   Andersson S. (1995) Molecular genetics of androgenic    17β-Hydroxysteroid Dehydrogenases. J. Steroid Biochem. Molec. Biol.,    55:533-534].-   Dong Y et al. (1998) “17β-hydroxysteroid dehydrogenases in human    bone cells” J. Bone Min. Res., 13:1539-1546-   WO 02/26706-   DE2411273-   Manhas M S, Sharma S D, Amin S G. (1972) “Heterocyclic compounds. 4.    Synthesis and antiinflammatory activity of some substituted    thienopyrimidinones” J Med. Chem. 15(1):106-7.-   Kapustina M V; Kharizomenova I A; Shvedov V I; Radkevich T P;    Shipilova L D (1992) “Synthesis and biological activity of    4,8-dioxo-3,4,5,6,7,8-hexahydrobenzothieno[2,3-d]pyrimidine    derivatives” Khimiko-Farmatsevticheskii Zhurnal 26(1):56-7-   Kapustina M V; Amelkin OYu; Kharizomenova I A; Shvedov V I; Filitis    L N (1991) “Synthesis and tuberculostatic activity of    benzothieno[2,3-d]pyrimidines” Khimiko-Farmatsevticheskii Zhurnal    25(7): 38-9-   Koffman B, Modarress K J, Beckerman T, Bashirelahi N. (1991)    “Evidence for involvement of tyrosine in estradiol binding by rat    uterus estrogen receptor.” J Steroid Biochem Mol. Biol. 38(2):135-9.-   WO 00/07996-   US 2003/0170292

1. A method of preparing a compound corresponding to formula (I)

wherein R₁ and R₂ represent the same or different alkyl, or one is alkyland the other is H, or R₁ and R₂ together with their binding sites forma cyclic 5-, 6-, 7- or 8-membered ring system, which is saturated orcontains one or more double bonds between the ring atoms, and which ringoptionally contains up to two heteroatoms in addition to the nitrogenatom where R₁ is attached, the number of N atoms being 0-2 and thenumber of O or S atoms each being 0-1, wherein said ring is optionallysubstituted with up to three substituents independently selected fromthe group consisting of alkyl, substituted alkyl, aryl, or arylalkyl,wherein the aryl group is optionally substituted, alkoxy, aryloxy,acyloxy, arylthio, alkylthio, arylsulfonyl, alkylsulfonyl, hydroxyl,oxo, halogen, amino, oxime, acyl, carboxyl, thiocarboxyl, and amido; R₃and R₄ form together with their binding sites a cyclic 5-, 6-, 7- or8-membered hydrocarbon ring system, which is saturated or contains oneor more double bonds between the carbon atoms, and wherein said ring isoptionally substituted with up to three substituents independentlyselected from the group consisting of alkyl, substituted alkyl, aryl orarylalkyl, wherein the aryl group is optionally substituted, alkoxy,aryloxy, acyloxy, arylthio, alkylthio, arylsulfonyl, alkylsulfonyl,hydroxyl, oxo, halogen, amino, oxime, acyl, carboxyl, thiocarboxyl, andamido; or a physiologically acceptable salt thereof; said methodcomprising: a) oxidizing a compound of formula 2

or a ring-substituted or ring-modified analogue thereof to give anoxo-substituted compound of formula 3 or an analogue thereof,

b) optionally further subjecting the oxo-substituted compound obtainedin a) to a Vilsmeier reaction to give a carbonylsubstituted compound offormula 4 or an analogue thereof,

c) optionally replacing the chlorosubstituent in the carbonylsubstitutedcompound obtained in b) by an alkylthio or an arylthio group bysubjecting to an appropriate thiol in the presence of a base to give anarylthio- or alkylthiosubstituted compound of formula 5 or an analoguethereof,

d) the arylthio- or alkylthiosubstituted compound obtained in c) isoptionally further i) reduced to a compound of formula 6,

ii) reacted with NH2OH to give a compound of formula 7,

Or e) the compound obtained in b) is optionally further i) reduced so asto replace the carbonyl group with hydroxyalkyl, or ii) subjected to anappropriate thiol in the presence of a base and acetone, so as toreplace the chloro substituent by a thiol group and to replace thecarbonyl group with an oxosubstituted alkenyl. or f) optionallysubjecting the compound obtained in a) to DMF acetal so as to introducea dimethylaminomethylene substituent in the ring next to the oxosubstituent.
 2. A method according to claim 1, wherein the oxidation instep a) is effected by subjecting the compound to PCC and celite.
 3. Amethod according to claim 1, wherein in b) the Vilsmeier reaction iseffected by POCl₃-DMF.
 4. A method of treating or inhibiting a steroidhormone dependent disease or disorder, said method comprisingadministering to a patient in need thereof an effective amount of acompound corresponding to formula (I)

wherein R₁ and R₂ represent the same or different alkyl, or one is alkyland the other is H; R₃ and R₄ form together with their binding sites acyclic 5-, 6-, 7- or 8-membered hydrocarbon ring system, which issaturated or contains one or more double bonds between the carbon atoms,and wherein said ring is optionally substituted with up to threesubstituents independently selected from the group consisting of alkyl,substituted alkyl, aryl or arylalkyl, wherein the aryl group isoptionally substituted, alkoxy, aryloxy, acyloxy, arylthio, alkylthio,arylsulfonyl, alkylsulfonyl, hydroxyl, oxo, halogen, amino, oxime, acyl,carboxyl, thiocarboxyl, and amido; or a physiologically acceptable saltthereof.
 5. A method according to claim 4, wherein said steroid hormonedependent disease or disorder is a steroid hormone dependent disease ordisorder requiring the inhibition of a 17β-hydroxysteroid dehydrogenaseenzyme.
 6. A method according to claim 5, wherein said steroid hormonedependent disease or disorder is a steroid hormone dependent disease ordisorder requiring the inhibition of the 17β-hydroxysteroiddehydrogenase type 1, 17β-hydroxysteroid dehydrogenase type 2 or17β-hydroxysteroid dehydrogenase type 3 enzyme.
 7. A method according toclaim 4, wherein said compound corresponds to formula (II)

wherein R₁ and R₂ represent the same or different C₁-C₈-alkyl, or one isC₁-C₈-alkyl and the other is H; the hydrocarbon chain—C(R5)-C(R6)-(CH)_(n)— of the ring-system adjacent the thiophene-ring issaturated or contains one or more double bonds between the carbon atoms;n is an integer from 1 to 4, and R5 and R6 are individually selectedfrom the group consisting of hydrogen, alkyl, substituted alkyl, aryl orarylalkyl, wherein the aryl group is optionally substituted, alkoxy,aryloxy, acyloxy, arylthio, alkylthio, arylsulfonyl, alkylsulfonyl,hydroxyl, oxo, halogen, amino, oxime, acyl, carboxyl, thiocarboxyl, andamido.
 8. A method according to claim 1, wherein the steroid hormonedependent disease or disorder is selected from the group consisting ofbreast cancer, prostate carcinoma, ovarian cancer, uterine cancer,endometrial cancer and endometrial hyperplasia, endometriosis, uterinefibroids, uterine leiomyoma, adenomyosis, dysmenorrhea, menorrhagia,metrorrhagia, prostadynia, benign prostatic hyperplasia, prostatitis,acne, seborrhea, hirsutism, androgenic alopecia, precocious puberty,adrenal hyperplasia, polycystic ovarian syndrome, urinary dysfunction,osteoporosis, multiple sclerosis, rheumatoid arthritis, Alzheimer'sdisease, colon cancer, tissue wounds, skin wrinkles and cataracts.